Laboratory of Developmental Biology, Disease Networks Researtch Unit, Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland.
Kvantum Institute, Infotech Oulu, University of Oulu, Oulu, Finland.
Dev Dyn. 2022 Mar;251(3):536-550. doi: 10.1002/dvdy.418. Epub 2021 Sep 16.
Tissue organoids derived from primary cells have high potential for studying organ development and diseases in numerous organs. They recreate the morphological structure and mimic the functions of given organ while being compact in size, easy to produce, and suitable for use in various experimental setups.
In this study we established the number of cells that form mouse kidney rudiments at E11.5, and generated renal organoids of various sizes from the mouse primary cells of the metanephric mesenchyme (MM). We investigated the ability of renal organoids to undergo nephrogenesis upon Wnt/ β-catenin pathway-mediated tubule induction with a GSK-3 inhibitor (BIO) or by initiation through the ureteric bud (UB). We found that 5000 cells of MM cells are necessary to successfully form renal organoids with well-structured nephrons as judged by fluorescent microscopy, transmission electron microscopy (TEM), and quantitative Polymerase Chain Reaction (qPCR). These mouse organoids also recapitulated renal secretion function in the proximal tubules.
We show that a significant decrease of cells used to generate renal mouse organoids in a dissociation/re-aggregation assay, does not interfere with development, and goes toward 3Rs. This enables generation of more experimental samples with one mouse litter, limiting the number of animals used for studies.
源自原代细胞的组织类器官在研究众多器官的器官发生和疾病方面具有很大的潜力。它们在体积小、易于生成且适合各种实验设置的同时，重现了给定器官的形态结构并模拟了其功能。
在这项研究中，我们确定了 E11.5 时形成小鼠肾脏原基的细胞数量，并从小鼠后肾间充质的原代细胞中生成了各种大小的肾类器官。我们研究了肾类器官在 Wnt/β-连环蛋白通路介导的小管诱导（用 GSK-3 抑制剂 BIO）或通过输尿管芽（UB）起始时进行肾发生的能力。我们发现，5000 个 MM 细胞足以成功形成具有荧光显微镜、透射电子显微镜（TEM）和定量聚合酶链反应（qPCR）判断的结构良好的肾单位的肾类器官。这些小鼠类器官还在近端小管中再现了肾脏分泌功能。
我们表明，在分离/再聚集测定中用于生成肾小鼠类器官的细胞数量显著减少，不会干扰发育，并且符合 3R 原则。这使得可以用一只小鼠的后代生成更多的实验样本，从而限制了用于研究的动物数量。
