Zimmer K P, le Coutre P, Aerts H M, Harzer K, Fukuda M, O'Brien J S, Naim H Y
Universitätskinderklinik, Westfälische Wilhelms-Universität Münster, D-48149 Münster, Germany.
J Pathol. 1999 Aug;188(4):407-14. doi: 10.1002/(SICI)1096-9896(199908)188:4<407::AID-PATH377>3.0.CO;2-Z.
Gaucher's disease (GD) is caused by an inherited deficiency of acid beta-glucosidase with storage of glucosylceramides in the lysosomes of macrophages. This study identifies a G202R mutation in the acid beta-glucosidase gene in an infant with severe neuronopathic (type 2) GD and only slightly reduced acid beta-glucosidase activity. Western blot analysis, pulse chase experiments, and the thin frozen section immunogold method were used to analyse the implications of this mutation on the pathogenesis, clinical heterogeneity and diagnostic evaluation of GD. The results show that acid beta-glucosidase persists in the patient's fibroblasts as a mannose-rich polypeptide in the endoplasmic reticulum and is not transported to the lysosomes. By contrast, high expression of the lysosome-associated membrane proteins LAMP-1 and LAMP-2, saposin C, and cathepsin D was observed in the patient's lysosomes. Immunogold labelling of the integral membrane proteins LAMP-1 and LAMP-2 increases significantly at the cell surface of Kupffer cells and fibroblasts as well as at the apical membrane of hepatocytes. In addition, LAMP-1 and LAMP-2 associate with the bilayer of stored glucosylceramide. It is concluded that defective intracellular transport of mutant acid beta-glucosidase from the endoplasmic reticulum to lysosomes leads to a more severe clinical phenotype than the residual enzyme activity may indicate. Furthermore, the detection of LAMP in the tubular bundles of undigested glucosylceramides, as well as their increased concentration at the surfaces of the affected cells, suggests that these proteins play a role in the storage or removal of substrate in GD. Intracellular targeting of acid beta-glucosidase and LAMP contributes to the broad phenotypic heterogeneity of GD.
戈谢病（GD）是由酸性β-葡萄糖苷酶遗传性缺乏导致葡糖神经酰胺在巨噬细胞溶酶体中蓄积引起的。本研究在一名患有严重神经病变型（2型）GD且酸性β-葡萄糖苷酶活性仅略有降低的婴儿中，鉴定出酸性β-葡萄糖苷酶基因中的一个G202R突变。采用蛋白质免疫印迹分析、脉冲追踪实验和薄冰冻切片免疫金法，分析该突变对GD发病机制、临床异质性和诊断评估的影响。结果显示，酸性β-葡萄糖苷酶在患者成纤维细胞中以内质网中富含甘露糖的多肽形式持续存在，未转运至溶酶体。相比之下，在患者溶酶体中观察到溶酶体相关膜蛋白LAMP-1和LAMP-2、鞘脂激活蛋白C和组织蛋白酶D的高表达。LAMP-1和LAMP-2这两种整合膜蛋白在库普弗细胞和成纤维细胞的细胞表面以及肝细胞的顶膜上的免疫金标记显著增加。此外，LAMP-1和LAMP-2与储存的葡糖神经酰胺双层结合。得出的结论是，突变的酸性β-葡萄糖苷酶从内质网到溶酶体的细胞内转运缺陷导致的临床表型比残余酶活性所提示的更为严重。此外，在未消化的葡糖神经酰胺管状束中检测到LAMP，以及它们在受影响细胞表面的浓度增加，表明这些蛋白在GD中底物的储存或清除中起作用。酸性β-葡萄糖苷酶和LAMP的细胞内靶向作用导致了GD广泛的表型异质性。
