Organoids derived from induced pluripotent stem (iPS) or embryonic stem (ES) cells have been evaluated as in vitro models of development and disease. However, maintaining these cells under long-term static culture conditions is difficult because of nutrition shortages and waste accumulation. To overcome these issues, perfusion culture systems are required for organoid technology. A system with a stable microenvironment, nutrient availability, and waste removal will accelerate organoid generation. The aim of this study was to develop a novel perfusion system for renal organoids by maintaining the air-liquid interface with a device fabricated using a 3D printer. Our results revealed slow flow at the organoid cultivation area based on microbead movement on the membrane, which depended on the perfusion rate under the membrane. Moreover, the perfused culture medium below the organoids via a porous membrane diffused throughout the organoids, maintaining the air-liquid interface. The diffusion rates within organoids were increased according to the flow rate of the culture medium under the membrane. The perfused culture medium also stimulated cytoskeletal and basement membrane re-organization associated with promotion tubular formation under 2.5 μL/min flow culture. In contrast, tubules in organoids were diminished at a flow rate of 10 μL/min. Our liquid-air interface perfusion system accelerated organization of the renal organoids. These results suggest that suitable perfusion conditions can accelerate organization of epithelial cells and tissues in renal organoids in vitro.

中文翻译：

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用气-液界面维持灌注培养以在体外刺激肾类器官中的上皮细胞组织。

源自诱导多能干 (iPS) 或胚胎干 (ES) 细胞的类器官已被评估为发育和疾病的体外模型。然而，由于营养短缺和废物积累，很难在长期静态培养条件下维持这些细胞。为了克服这些问题，类器官技术需要灌注培养系统。具有稳定微环境、养分可用性和废物清除的系统将加速类器官的产生。本研究的目的是通过使用 3D 打印机制造的设备维持气液界面，开发一种用于肾类器官的新型灌注系统。我们的结果表明，基于膜上微珠运动的类器官培养区域的缓慢流动，这取决于膜下的灌注速率。而且，通过在整个类器官中扩散的多孔膜，在类器官下方灌注培养基，保持气液界面。类器官内的扩散速率根据膜下培养基的流速增加。在 2.5 μL/min 流式培养下，灌注培养基还刺激了与促进管状形成相关的细胞骨架和基底膜重组。相反，类器官中的小管以 10 μL/min 的流速减少。我们的液-气界面灌注系统加速了肾脏类器官的组织。这些结果表明，合适的灌注条件可以加速体外肾类器官中上皮细胞和组织的组织。保持气液界面。类器官内的扩散速率根据膜下培养基的流速增加。在 2.5 μL/min 流式培养下，灌注培养基还刺激了与促进管状形成相关的细胞骨架和基底膜重组。相反，类器官中的小管以 10 μL/min 的流速减少。我们的液-气界面灌注系统加速了肾脏类器官的组织。这些结果表明，合适的灌注条件可以加速体外肾类器官中上皮细胞和组织的组织。保持气液界面。类器官内的扩散速率根据膜下培养基的流速增加。在 2.5 μL/min 流式培养下，灌注培养基还刺激了与促进管状形成相关的细胞骨架和基底膜重组。相反，类器官中的小管以 10 μL/min 的流速减少。我们的液-气界面灌注系统加速了肾脏类器官的组织。这些结果表明，合适的灌注条件可以加速体外肾类器官中上皮细胞和组织的组织。在 2.5 μL/min 流式培养下，灌注培养基还刺激了与促进管状形成相关的细胞骨架和基底膜重组。相反，类器官中的小管以 10 μL/min 的流速减少。我们的液-气界面灌注系统加速了肾脏类器官的组织。这些结果表明，合适的灌注条件可以加速体外肾类器官中上皮细胞和组织的组织。在 2.5 μL/min 流式培养下，灌注培养基还刺激了与促进管状形成相关的细胞骨架和基底膜重组。相反，类器官中的小管以 10 μL/min 的流速减少。我们的液-气界面灌注系统加速了肾脏类器官的组织。这些结果表明，合适的灌注条件可以加速体外肾类器官中上皮细胞和组织的组织。