Grimm Marcus O W, Hundsdörfer Benjamin, Grösgen Sven, Mett Janine, Zimmer Valerie C, Stahlmann Christoph P, Haupenthal Viola J, Rothhaar Tatjana L, Lehmann Johannes, Pätzold Andreas, Zinser Eva G, Tanila Heikki, Shen Jie, Müller Ulrike, Grimm Heike S, Hartmann Tobias
Experimental Neurology, Saarland University, Homburg/Saar, Germany.
Cell Physiol Biochem. 2014;34(1):92-110. doi: 10.1159/000362987. Epub 2014 Jun 16.
Gangliosides were found to be associated with Alzheimer's disease (AD). Here we addressed a potential function of γ-secretase (presenilin) dependent cleavage of the amyloid-precursor-protein (APP) in the regulation of ganglioside de novo synthesis.
To identify a potential role of γ-secretase and APP in ganglioside de novo synthesis we used presenilin (PS) deficient and APP deficient cells and mouse brains, mutated PS as well as transgenic mice and AD post mortem brains. Changes in glucosylceramide synthase (GCS) activity were identified by incorporation of radiolabeled UDP-glucose in glucosylceramide, changes in gene expression via real-time PCR and Western blot analysis. Alterations in ganglioside levels were determined by thin layer chromatography and mass spectrometry.
We found that PS and APP deficiency, in vitro and in vivo, resulted in increased GCS gene expression, elevated enzyme activity and thus increased glucosylceramide and total ganglioside level. Using a specific γ-secretase inhibitor revealed that PS proteolytic activity alters ganglioside homeostasis. By the use of mutated PS causing early onset AD in cell culture and transgenic mice we found that GCS is increased in AD, further substantiated by the use of AD post mortem brains, suffering from sporadic AD.
APP processing regulates ganglioside de novo synthesis and is affected in AD.
已发现神经节苷脂与阿尔茨海默病（AD）相关。在此，我们探讨了淀粉样前体蛋白（APP）经γ-分泌酶（早老素）依赖性切割在神经节苷脂从头合成调节中的潜在作用。
为确定γ-分泌酶和APP在神经节苷脂从头合成中的潜在作用，我们使用了早老素（PS）缺陷型和APP缺陷型细胞及小鼠脑、突变型PS以及转基因小鼠和AD尸检脑。通过将放射性标记的UDP-葡萄糖掺入葡糖神经酰胺来鉴定葡糖神经酰胺合酶（GCS）活性的变化，通过实时PCR和蛋白质印迹分析来鉴定基因表达的变化。通过薄层色谱法和质谱法测定神经节苷脂水平的改变。
我们发现，在体外和体内，PS和APP缺陷均导致GCS基因表达增加、酶活性升高，从而使葡糖神经酰胺和总神经节苷脂水平升高。使用特异性γ-分泌酶抑制剂表明，PS的蛋白水解活性会改变神经节苷脂的稳态。通过使用在细胞培养和转基因小鼠中导致早发性AD的突变型PS，我们发现AD中GCS增加，使用散发性AD的AD尸检脑进一步证实了这一点。
APP加工调节神经节苷脂从头合成，且在AD中受到影响。
