淀粉样β(Aβ)肽在阿尔茨海默病(AD)病理中具有重要作用。肽长度可以在37和49个氨基酸之间变化。Aβ1-42被认为是与疾病相关的长度。然而,Aβ1-40也存在于Aβ斑块中,并且已显示与Aβ1-42形成交织的原纤维。这些肽以前也显示形成不同的原纤维构象,建议与疾病表型有关。进行更有代表性的体外实验,揭示不同原纤维构象对神经元的影响至关重要。因此,我们以100:0、90:10、75:25、50:50、25:75、10:90和0:100的浓度将不同的Aβ1-40:42比例原纤维化24小时(早期原纤维)或7天(老化原纤维)。然后根据原纤维宽度对其进行表征,LCO染色和抗体染色。我们进一步用不同的原纤维攻击分化的神经元样SH-SY5Y人细胞,并测量Aβ含量,在三个不同的时间点:3、24和72h的细胞毒性和自噬功能。我们的结果表明,Aβ1-40:42比例和原纤维成熟都会影响原纤维的构象。我们进一步显示了这些构象变化对常用Aβ抗体亲和力的影响,主要影响富含Aβ1-40的聚集体。此外,我们证明了神经元分化的人类细胞对聚集体的吸收,其中具有较高Aβ1-42比率的聚集体通常引起较高的Aβ细胞水平。Aβ丰度的这些差异不会引起细胞毒性或自噬激活的变化。我们的结果表明,考虑Aβ原纤维的构象差异的重要性,因为这可能对Aβ抗体检测产生根本性影响。总的来说,这些见解强调了需要进一步探索构象不同原纤维的影响,以及需要可靠地产生与疾病相关的Aβ聚集体.
The amyloid β (Aβ) peptide has a central role in Alzheimer\'s disease (AD) pathology. The peptide length can vary between 37 and 49 amino acids, with Aβ1-42 being considered the most disease-related length. However, Aβ1-40 is also found in Aβ plaques and has shown to form intertwined fibrils with Aβ1-42. The peptides have previously also shown to form different fibril conformations, proposed to be related to disease phenotype. To conduct more representative in vitro experiments, it is vital to uncover the impact of different fibril conformations on neurons. Hence, we fibrillized different Aβ1-40:42 ratios in concentrations of 100:0, 90:10, 75:25, 50:50, 25:75, 10:90 and 0:100 for either 24 h (early fibrils) or 7 days (aged fibrils). These were then characterized based on fibril width, LCO-staining and antibody-staining. We further challenged differentiated neuronal-like SH-SY5Y human cells with the different fibrils and measured Aβ content, cytotoxicity and autophagy function at three different time-points: 3, 24, and 72 h. Our results revealed that both Aβ1-40:42 ratio and fibril maturation affect
conformation of fibrils. We further show the impact of these
conformation changes on the affinity to commonly used Aβ antibodies, primarily affecting Aβ1-40 rich aggregates. In addition, we demonstrate uptake of the aggregates by neuronally differentiated human cells, where aggregates with higher Aβ1-42 ratios generally caused higher cellular levels of Aβ. These differences in Aβ abundance did not cause changes in cytotoxicity nor in autophagy activation. Our results show the importance to consider conformational differences of Aβ fibrils, as this can have fundamental impact on Aβ antibody detection. Overall, these insights underline the need for further exploration of the impact of conformationally different fibrils and the need to reliably produce disease relevant Aβ aggregates.