Department of Pediatrics, University Hospitals Leuven Campus Gasthuisberg, B-3000 Leuven, Belgium.
Laboratory of Pediatric Nephrology, Department of Development & Regeneration, KU Leuven Campus Gasthuisberg, B-3000 Leuven, Belgium.
Cells. 2022 Apr 6;11(7):1245. doi: 10.3390/cells11071245.
Nephropathic cystinosis is an inherited lysosomal storage disorder caused by pathogenic variants in the cystinosin () gene and is characterized by the excessive shedding of proximal tubular epithelial cells (PTECs) and podocytes into urine, development of the renal Fanconi syndrome and end-stage kidney disease (ESKD). We hypothesized that in compensation for epithelial cell losses, cystinosis kidneys undertake a regenerative effort, and searched for the presence of kidney progenitor cells (KPCs) in the urine of cystinosis patients. Urine was cultured in a specific progenitor medium to isolate undifferentiated cells. Of these, clones were characterized by qPCR, subjected to a differentiation protocol to PTECs and podocytes and assessed by qPCR, Western blot, immunostainings and functional assays. Cystinosis patients voided high numbers of undifferentiated cells in urine, of which various clonal cell lines showed a high capacity for self-renewal and expressed kidney progenitor markers, which therefore were assigned as cystinosis urine-derived KPCs (Cys-uKPCs). Cys-uKPC clones showed the capacity to differentiate between functional PTECs and/or podocytes. Gene addition with wild-type using lentiviral vector technology resulted in significant reductions in cystine levels. We conclude that KPCs present in the urine of cystinosis patients can be isolated, differentiated and complemented with in vitro, serving as a novel tool for disease modeling.
遗传性胱氨酸贮积症是一种溶酶体贮积病，由胱氨酸转运蛋白（）基因的致病性变异引起，其特征是近端肾小管上皮细胞（PTEC）和足细胞过度脱落到尿液中，导致范可尼综合征和终末期肾病（ESKD）。我们假设，为了代偿上皮细胞的丢失，胱氨酸贮积症的肾脏会进行再生努力，因此我们在胱氨酸贮积症患者的尿液中寻找肾脏祖细胞（KPC）的存在。将尿液在特定的祖细胞培养基中培养以分离未分化细胞。对这些细胞进行 qPCR 特征分析，然后进行向 PTEC 和足细胞的分化方案，并通过 qPCR、Western blot、免疫染色和功能测定进行评估。胱氨酸贮积症患者的尿液中大量排泄未分化细胞，其中各种克隆细胞系具有很强的自我更新能力，并表达肾脏祖细胞标记物，因此被鉴定为胱氨酸贮积症尿液衍生的 KPC（Cys-uKPC）。Cys-uKPC 克隆显示出分化为功能性 PTEC 和/或足细胞的能力。使用慢病毒载体技术进行野生型的基因添加导致胱氨酸水平显著降低。我们的结论是，胱氨酸贮积症患者尿液中的 KPC 可以被分离、分化，并通过体外补充野生型进行补充，为疾病建模提供了一种新的工具。
