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Abstract：
The neuronal ceroid lipofuscinoses (NCLs, Batten disease) are inherited neurodegenerative lysosomal storage diseases caused by mutations in several different genes. Mutations in CLN5 cause a variant late-infantile human disease and some cases of juvenile and adult clinical disease. NCLs also occur in animals, and a flock of New Zealand Borderdale sheep with a CLN5 splice-site mutation has been developed for model studies. Dissociated mixed neural cells from CLN5-deficient foetal sheep brains contained no obvious storage bodies at plating but these accumulated rapidly in culture, mainly in microglial cells and also in neurons and astrocytes. Accumulation was very obvious after a week, as monitored by fluorescent microscopy and immunostaining for subunit c of mitochondrial ATP synthase. Photography at intervals revealed the dynamic nature of the cultures and a flow of storage bodies between cells, specifically the phagocytosis of storage-body containing cells by microglia and incorporation of the storage bodies into the host cells. No storage was observed in cultured control cells. Transduction of cell cultures with a lentiviral vector expressing a C-terminal Myc tagged CLN5 resulted in secretion of post-translationally glycosylated and processed CLN5. Transduction of CLN5-deficient cultures with this construct rapidly reversed storage body accumulation, to less than half in only six days. These results show that storage body accumulation is reversible with enzyme correction and support the use of these cultures for testing of therapeutics prior to whole animal studies.
摘要：
神经元类脂褐斑病(NCLs，Batten病）是由几种不同基因的突变引起的遗传性神经退行性溶酶体贮积病。CLN5中的突变会导致婴儿后期人类疾病以及一些青少年和成人临床疾病病例。NCL也发生在动物中，并且已开发出具有CLN5剪接位点突变的新西兰Borderdale羊群用于模型研究。从缺乏CLN5的胎儿绵羊大脑中分离的混合神经细胞在平板上没有明显的储存体，但这些细胞在培养物中迅速积累，主要存在于小胶质细胞中，也存在于神经元和星形胶质细胞中。一周后积累非常明显，通过荧光显微镜和免疫染色监测线粒体ATP合酶的亚基c。间隔摄影揭示了培养物的动态性质和细胞之间的储存体流动，特别是小胶质细胞吞噬含有储存体的细胞，并将储存体掺入宿主细胞。在培养的对照细胞中未观察到储存。用表达C末端Myc标记的CLN5的慢病毒载体转导细胞培养物导致翻译后糖基化和加工的CLN5的分泌。用这种构建物转导缺乏CLN5的培养物迅速逆转了储存体的积累，在六天内不到一半。这些结果表明，通过酶校正，储存体的积累是可逆的，并且支持在整个动物研究之前将这些培养物用于测试治疗剂。
