Williams James, Hurling Chloe, Munir Sabrina, Harley Peter, Machado Carolina Barcellos, Cujba Ana-Maria, Alvarez-Fallas Mario, Danovi Davide, Lieberam Ivo, Sancho Rocio, Beales Philip, Watt Fiona M
Centre for Gene Therapy and Regenerative Medicine, King's College London, Guy's Hospital, London, United Kingdom.
Bit.bio, Babraham Research Campus, Cambridge, United Kingdom.
Front Cell Dev Biol. 2023 Jun 2;11:1163825. doi: 10.3389/fcell.2023.1163825. eCollection 2023.
Bardet-Biedl syndrome (BBS) is a ciliopathy with pleiotropic effects on multiple tissues, including the kidney. Here we have compared renal differentiation of iPS cells from healthy and BBS donors. High content image analysis of WT1-expressing kidney progenitors showed that cell proliferation, differentiation and cell shape were similar in healthy, , , and mutant lines. We then examined three patient lines with mutations in a 3D kidney organoid system. The line with the most deleterious mutation, with low BBS10 expression, expressed kidney marker genes but failed to generate 3D organoids. The other two patient lines expressed near normal levels of mRNA and generated multiple kidney lineages within organoids when examined at day 20 of organoid differentiation. However, on prolonged culture (day 27) the proximal tubule compartment degenerated. Introducing wild type into the most severely affected patient line restored organoid formation, whereas CRISPR-mediated generation of a truncating BBS10 mutation in a healthy line resulted in failure to generate organoids. Our findings provide a basis for further mechanistic studies of the role of BBS10 in the kidney.
巴德-比埃尔综合征（BBS）是一种对包括肾脏在内的多种组织具有多效性影响的纤毛病。在此，我们比较了来自健康供体和BBS供体的诱导多能干细胞（iPS细胞）的肾脏分化情况。对表达WT1的肾脏祖细胞进行的高内涵图像分析表明，健康、 、 及 突变系中的细胞增殖、分化和细胞形态相似。然后，我们在三维肾脏类器官系统中检测了三个携带 突变的患者系。具有最有害突变且BBS10表达水平低的细胞系表达了肾脏标记基因，但未能生成三维类器官。另外两个患者系在类器官分化第20天时检测，其 信使核糖核酸表达接近正常水平，并在类器官内生成了多种肾脏谱系。然而，在长期培养（第27天）时，近端小管区发生了退化。将野生型 导入受影响最严重的患者细胞系可恢复类器官形成，而在健康细胞系中通过CRISPR介导产生截短的BBS10突变则导致无法生成类器官。我们的研究结果为进一步深入研究BBS10在肾脏中的作用机制提供了基础。
