Department of Biomedical and Clinical Sciences and Department of Clinical Pathology, Linköping University, Linköping, Sweden.
Department of Physics, Chemistry and Biology, Linköping University, Linköping, Sweden.
J Neurochem. 2024 Sep;168(9):3308-3322. doi: 10.1111/jnc.16201. Epub 2024 Aug 12.
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.
淀粉样蛋白 β（Aβ）肽在阿尔茨海默病（AD）病理学中起着核心作用。该肽的长度在 37 到 49 个氨基酸之间变化，其中 Aβ1-42 被认为与疾病的相关性最强。然而，Aβ1-40 也存在于 Aβ 斑块中，并已显示与 Aβ1-42 形成交织的原纤维。这些肽以前也表现出形成不同的原纤维构象，这被认为与疾病表型有关。为了进行更具代表性的体外实验，揭示不同原纤维构象对神经元的影响至关重要。因此，我们将不同浓度的 Aβ1-40:42 比值（100:0、90:10、75:25、50:50、25:75、10:90 和 0:100）分别在 24 小时（早期原纤维）或 7 天（老化原纤维）的时间点进行纤维化。然后根据原纤维宽度、LCO 染色和抗体染色对其进行特征分析。我们进一步用不同的原纤维挑战分化的神经元样 SH-SY5Y 人细胞，并在三个不同的时间点（3、24 和 72 小时）测量 Aβ 含量、细胞毒性和自噬功能。我们的结果表明，Aβ1-40:42 比值和原纤维成熟度都影响原纤维的构象。我们进一步表明，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 的聚集物。此外，我们证明了神经元分化的人类细胞对聚集物的摄取，其中具有更高 Aβ1-42 比值的聚集物通常会导致细胞内 Aβ 水平更高。这些 Aβ 丰度的差异不会导致细胞毒性或自噬激活的变化。我们的结果表明，考虑 Aβ 原纤维的构象差异非常重要，因为这可能对 Aβ 抗体检测产生根本影响。总的来说，这些结果强调了需要进一步探索构象不同的原纤维的影响，以及需要可靠地产生与疾病相关的 Aβ 聚集物的必要性。
