Drabbe Emma, Pelaez Daniel, Agarwal Ashutosh
Department of Biomedical Engineering, University of Miami, Coral Gables, FL, USA.
Bascom Palmer Eye Institute, Department of Ophthalmology, University of Miami Miller School of Medicine, 1638 NW 10th Ave., Miami, FL 33136, USA.
Lab Chip. 2025 Mar 25;25(7):1626-1636. doi: 10.1039/d4lc00771a.
An oxygen gradient across the retina plays a crucial role in its development and function. The inner retina resides in a hypoxic environment (2% O) adjacent to the vitreous cavity. Oxygenation levels rapidly increase towards the outer retina (18% O) at the choroid. In addition to retinal stratification, oxygen levels are critical for the health of retinal ganglion cells (RGCs), which relay visual information from the retina to the brain. Human stem cell derived retinal organoids are being engineered to mimic the structure and function of human retina for applications such as disease modeling, development of therapeutics, and cell replacement therapies. However, rapid degeneration of the retinal ganglion cell layers are a common limitation of human retinal organoid platforms. We report the design of a novel retinal organoid chip (ROC) that maintains a physiologically relevant oxygen gradient and allows the maturation of inner and outer retinal cell phenotypes for human retinal organoids. Our PDMS-free ROC holds 55 individual retinal organoids that were manually seeded, cultured for extended periods (over 150 days), imaged , and retrieved. ROC was designed from first principles of liquid and gas mass transport, and fabricated from biologically- and chemically inert materials using rapid prototyping techniques such as micromachining, laser cutting, 3D printing and bonding. After computational and experimental validation of oxygen gradients, human induced pluripotent stem cell derived retinal organoids were transferred into the ROC, differentiated, cultured and imaged within the chip. ROCs that maintained active perfusion and stable oxygen gradients were successful in inducing higher viability of RGCs within retinal organoids than static controls, or ROC without oxygen gradients. Our physiologically relevant and higher-throughput retinal organoid culture system is well suited for applications in studying developmental perturbations to primate retinogenesis, including those driven by inherited traits, fetal environmental exposure to toxic agents, or acquired by genetic mutations, such as retinoblastoma.
视网膜上的氧梯度在其发育和功能中起着至关重要的作用。视网膜内层位于与玻璃体腔相邻的低氧环境（2%氧气）中。向脉络膜方向，外层视网膜的氧合水平迅速升高（18%氧气）。除了视网膜分层外，氧水平对于视网膜神经节细胞（RGCs）的健康也至关重要，这些细胞将视觉信息从视网膜传递到大脑。人类干细胞衍生的视网膜类器官正在被设计用来模拟人类视网膜的结构和功能，以用于疾病建模、治疗方法开发和细胞替代疗法等应用。然而，视网膜神经节细胞层的快速退化是人类视网膜类器官平台的一个常见限制。我们报告了一种新型视网膜类器官芯片（ROC）的设计，该芯片能维持生理相关的氧梯度，并允许人类视网膜类器官的内层和外层视网膜细胞表型成熟。我们不含聚二甲基硅氧烷（PDMS）的ROC容纳55个单独的视网膜类器官，这些类器官是手动接种的，经过长时间（超过150天）培养、成像并取出。ROC是根据液体和气体质量传输的基本原理设计的，并使用微加工、激光切割、3D打印和键合等快速成型技术，由生物和化学惰性材料制成。在对氧梯度进行计算和实验验证后，将人类诱导多能干细胞衍生的视网膜类器官转移到ROC中，在芯片内进行分化、培养和成像。与静态对照或无氧梯度的ROC相比，维持主动灌注和稳定氧梯度的ROC成功地诱导了视网膜类器官内RGCs的更高存活率。我们生理相关且高通量的视网膜类器官培养系统非常适合用于研究灵长类视网膜发生的发育扰动，包括那些由遗传特征、胎儿环境暴露于有毒物质或因基因突变（如视网膜母细胞瘤）而获得的扰动。
