Nilselid A-M, Davidsson Pia, Nägga Katarina, Andreasen Niels, Fredman Pam, Blennow Kaj
Institute of Clinical Neuroscience, The Sahlgrenska Academy at Göteborg University, Sahlgrenska University Hospital, SE 431 80 Mölndal, Sweden.
Neurochem Int. 2006 Jun;48(8):718-28. doi: 10.1016/j.neuint.2005.12.005. Epub 2006 Feb 21.
Clusterin is suggested to be involved in the pathogenesis of Alzheimer's disease. Clusterin expression is increased in brain tissue in affected regions of Alzheimer patients, and intense clusterin staining is found in both senile plaques and in neuronal and glia cells. In contrast, the cerebrospinal fluid level of clusterin in Alzheimer patients has, thus far, been found unchanged. Clusterin is a glycosylated protein, and an alteration of its glycosylation in Alzheimer's disease might influence accurate quantification in cerebrospinal fluid through interference of antibody binding to the protein. Using enzymatic deglycosylation of clusterin isolated from cerebrospinal fluid, we found that the carbohydrates attached to clusterin were of the N-linked type and sialic acids. Based on this finding, cerebrospinal fluid samples from Alzheimer patients (n=99) and controls (n=39) were analysed. The samples were treated with peptide: N-glycanase F, cleaving off N-linked carbohydrates, and clusterin was quantified before and after deglycosylation using a new sandwich enzyme-linked immunosorbent assay. Clusterin was significantly increased in Alzheimer patients, in both native (7.17+/-2.43 AU versus 5.73+/-2.09 AU; p=0.002), and deglycosylated samples (12.19+/-5.00 AU versus 9.68+/-4.38 AU; p=0.004). Deglycosylation led to increased measured levels of clusterin by 70% (p<0.001) in Alzheimer patients and 67% (p<0.001) in controls. These findings indicate that glycosylation of proteins may interfere with their quantification. The results show that clusterin is significantly increased in cerebrospinal fluid from Alzheimer patients as a group, supporting that clusterin might be involved in the pathogenesis of Alzheimer's disease. However, the individual clusterin levels overlap between the two groups, and thus cerebrospinal fluid clusterin measurement is not suitable as a biochemical marker in the diagnosis of Alzheimer's disease.
有研究表明，簇集素与阿尔茨海默病的发病机制有关。在阿尔茨海默病患者受影响区域的脑组织中，簇集素表达增加，并且在老年斑以及神经元和神经胶质细胞中均发现强烈的簇集素染色。相比之下，迄今为止发现阿尔茨海默病患者脑脊液中的簇集素水平没有变化。簇集素是一种糖基化蛋白，其在阿尔茨海默病中的糖基化改变可能会通过干扰抗体与该蛋白的结合而影响脑脊液中的准确定量。通过对从脑脊液中分离出的簇集素进行酶促去糖基化处理，我们发现与簇集素相连的碳水化合物为N-连接型且含有唾液酸。基于这一发现，对阿尔茨海默病患者（n = 99）和对照组（n = 39）的脑脊液样本进行了分析。样本用肽：N-聚糖酶F处理，切断N-连接的碳水化合物，并使用一种新的夹心酶联免疫吸附测定法在去糖基化前后对簇集素进行定量。在阿尔茨海默病患者中，无论是天然样本（7.17±2.43 AU对5.73±2.09 AU；p = 0.002）还是去糖基化样本（12.19±5.00 AU对9.68±4.38 AU；p = 0.004），簇集素均显著增加。去糖基化导致阿尔茨海默病患者中簇集素的测量水平增加70%（p < 0.001），对照组增加67%（p < 0.001）。这些发现表明蛋白质的糖基化可能会干扰其定量。结果表明，作为一个群体，阿尔茨海默病患者脑脊液中的簇集素显著增加，支持簇集素可能参与阿尔茨海默病发病机制的观点。然而，两组之间个体的簇集素水平存在重叠，因此脑脊液簇集素测量不适合作为阿尔茨海默病诊断的生化标志物。
