ABSTRACT
Studies on the mechanisms of neuronal amyloid-ß (Aß) internalisation are crucial for understanding the neuropathological progression of Alzheimer's disease (AD). We here investigated how extracellular Aß peptides are internalised and focused on three different pathways: (i) via endocytic mechanisms, (ii) via the receptor for advanced glycation end products (RAGE) and (iii) via the pore-forming protein perforin. Both Aß40 and Aß42 were internalised in retinoic acid differentiated neuroblastoma (RA-SH-SY5Y) cells. A higher concentration was required for Aß40 (250 nM) compared with Aß42 (100 nM). The internalised Aß40 showed a dot-like pattern of distribution whereas Aß42 accumulated in larger and distinct formations. By confocal microscopy, we showed that Aß40 and Aß42 co-localised with mitochondria, endoplasmic reticulum (ER) and lysosomes. Aß treatment of human primary cortical neurons (hPCN) confirmed our findings in RA-SH-SY5Y cells, but hPCN were less sensitive to Aß; therefore, a 20 (Aß40) and 50 (Aß42) times higher concentration was needed for inducing uptake. The blocking of endocytosis completely inhibited the internalisation of Aß peptides in RA-SH-SY5Y cells and hPCN, indicating that this is a major pathway by which Aß enters the cells. In addition, the internalisation of Aß42, but not Aß40, was reduced by 55 % by blocking RAGE. Finally, for the first time we showed that pore formation in cell membranes by perforin led to Aß internalisation in hPCN. Understanding how Aß is internalised sheds light on the pathological role of Aß and provides further ideas of inhibitory strategies for preventing Aß internalisation and the spreading of neurodegeneration in AD.
