Cell Biology Unit, de Duve Institute and Université Catholique de Louvain, Brussels, Belgium;
Inserm, U574, Hôpital Necker-Enfants Malades and Université Paris Descartes, Sorbonne Paris Cité, Institut Imagine, Paris, France;
J Am Soc Nephrol. 2014 Jun;25(6):1256-69. doi: 10.1681/ASN.2013060598. Epub 2014 Feb 13.
Cystinosis, a main cause of Fanconi syndrome, is reproduced in congenic C57BL/6 cystinosin knockout (KO) mice. To identify the sequence of pathogenic and adaptation mechanisms of nephropathic cystinosis, we defined the onset of Fanconi syndrome in KO mice between 3 and 6 months of age and analyzed the correlation with structural and functional changes in proximal tubular cells (PTCs), with focus on endocytosis of ultrafiltrated disulfide-rich proteins as a key source of cystine. Despite considerable variation between mice at the same age, typical event sequences were delineated. At the cellular level, amorphous lysosomal inclusions preceded cystine crystals and eventual atrophy without crystals. At the nephron level, lesions started at the glomerulotubular junction and then extended distally. In situ hybridization and immunofluorescence revealed progressive loss of expression of megalin, cubilin, sodium-glucose cotransporter 2, and type IIa sodium-dependent phosphate cotransporter, suggesting apical dedifferentiation accounting for Fanconi syndrome before atrophy. Injection of labeled proteins revealed that defective endocytosis in S1 PTCs led to partial compensatory uptake by S3 PTCs, suggesting displacement of endocytic load and injury by disulfide-rich cargo. Increased PTC apoptosis allowed luminal shedding of cystine crystals and was partially compensated for by tubular proliferation. We conclude that lysosomal storage triggered by soluble cystine accumulation induces apical PTC dedifferentiation, which causes transfer of the harmful load of disulfide-rich proteins to more distal cells, possibly explaining longitudinal progression of swan-neck lesions. Furthermore, our results suggest that subsequent adaptation mechanisms include lysosomal clearance of free and crystalline cystine into urine and ongoing tissue repair.
胱氨酸贮积症是范可尼综合征的主要病因之一，可在同基因 C57BL/6 胱氨酸溶酶体失活（KO）小鼠中复制。为了确定胱氨酸贮积症肾病患者的致病和适应机制的先后顺序，我们确定 KO 小鼠范可尼综合征的发病时间在 3 至 6 月龄之间，并分析其与近端肾小管细胞（PTC）结构和功能变化的相关性，重点是将富含二硫键的超滤液蛋白的内吞作用作为胱氨酸的主要来源。尽管同一年龄的小鼠之间存在相当大的差异，但仍可以描绘出典型的事件序列。在细胞水平上，无定形溶酶体包涵体先于胱氨酸晶体出现，最终出现萎缩但无晶体。在肾单位水平上，病变从肾小球-肾小管交界处开始，然后向远端扩展。原位杂交和免疫荧光显示，多配体蛋白聚糖、内因子-cubilin 受体、钠-葡萄糖共转运蛋白 2 和 IIa 型钠依赖性磷酸盐共转运蛋白的表达逐渐丧失，提示顶泌细胞去分化导致萎缩前出现范可尼综合征。标记蛋白的注射显示 S1 PTC 的内吞缺陷导致 S3 PTC 的部分代偿性摄取，表明富含二硫键的货物导致内吞负荷和损伤的转移和替代。PTC 凋亡增加导致胱氨酸晶体从管腔脱落，并部分由管状增殖补偿。我们得出结论，溶酶体储存被可溶性胱氨酸积累触发，导致顶泌 PTC 去分化，从而将富含二硫键的蛋白质的有害负荷转移到更远端的细胞，这可能解释了天鹅颈样病变的纵向进展。此外，我们的研究结果表明，随后的适应机制包括将游离和结晶胱氨酸从细胞内溶酶体清除到尿液中，并进行持续的组织修复。
