P4–070: Amyloid precursor–like proteins 1 and 2 influence proteolytic processing of the amyloid precursor protein by alpha– and beta–secretase

uptake system in TR-BBB cells, an immortalized cell line derived from brain capillary endothelial cells of temperature-sensitive large T antigen transgenic rat, that recapitulates many of the characteristics of BBB. Results: TR-BBB cells showed a rapid internalization of [125I]-A 1-40, which is saturated at 10 min. Pulse-chase analysis demonstrated that A , once internalized, is rapidly effluxed into the culture media, suggesting that A receptor can be recycled onto the plasma membrane. This uptake was temperature-sensitive and blocked by excess amounts of unlabeled A 1-40 or A 1-42 in a concentration-dependent fashion. These data indicated that TR-BBB cells internalize A by a receptor-mediated mechanism and that A 1-40 and A 1-42 share the same receptor. We also found that receptorassociated protein (RAP), an antagonist of LDL receptor family proteins, significantly inhibits the binding and uptake of A in TR-BBB cells by 80%. We also found that apolipoprotein E (apoE) also inhibited the uptake of A . However, both MEF-1 cells (expressing LRP1 endogenously) and PEA13 cells (LRP1 homozygous deficient) did not internalize A . An in vitro binding assay using immobilized LRP1 also suggested that LRP1 does not directly bind A . Conclusions: These data indicated that a RAP-sensitive, certain LDL receptor family member mediates A uptake in TR-BBB cells, although it still remains possible that LRP1 indirectly mediates the uptake of A through an as yet unknown co-receptor system.