Department of Developmental Biology, University of Pittsburgh, School of Medicine;
Department of Developmental Biology, University of Pittsburgh, School of Medicine.
J Vis Exp. 2021 Apr 13(170). doi: 10.3791/62452.
Kidney organoids generated from hPSCs have provided an unlimited source of renal tissue. Human kidney organoids are an invaluable tool for studying kidney disease and injury, developing cell-based therapies, and testing new therapeutics. For such applications, large numbers of uniform organoids and highly reproducible assays are needed. We have built upon our previously published kidney organoid protocol to improve the overall health of the organoids. This simple, robust 3D protocol involves the formation of uniform embryoid bodies in minimum component medium containing lipids, insulin-transferrin-selenium-ethanolamine supplement and polyvinyl alcohol with GSK3 inhibitor (CHIR99021) for 3 days, followed by culture in knock-out serum replacement (KOSR)-containing medium. In addition, agitating assays allows for reduction in clumping of the embryoid bodies and maintaining a uniform size, which is important for reducing variability between organoids. Overall, the protocol provides a fast, efficient, and cost-effective method for generating large quantities of kidney organoids.
由 hPSC 生成的肾类器官为肾脏组织提供了无限的来源。人类肾类器官是研究肾脏疾病和损伤、开发基于细胞的疗法以及测试新疗法的宝贵工具。对于此类应用，需要大量均匀的类器官和高度可重复的测定。我们在之前发表的肾类器官方案的基础上进行了改进，以提高类器官的整体健康水平。该简单、稳健的 3D 方案涉及在含有脂质、胰岛素转铁蛋白-硒-乙醇胺补充剂和聚乙烯醇以及 GSK3 抑制剂（CHIR99021）的最小成分培养基中形成均匀的胚状体，培养 3 天，然后在含有无血清替代物（KOSR）的培养基中培养。此外，搅拌测定可减少胚状体的结块并保持均匀的大小，这对于减少类器官之间的变异性很重要。总体而言，该方案提供了一种快速、高效且具有成本效益的方法来生成大量的肾类器官。
