Periodic Report Summary - ADPROGRES (Alzheimer disease progression: Molecular studies of Abeta amyloid peptides aggregation and trafficking in neuronal cells) Alzheimer's disease (AD) is characterized by the accumulation of amyloid beta (Abeta) peptides in amyloid deposits in the cerebral tissue. Many reports found strong evidence that intermediates in the aggregation process call 'oligomers' are the principal pathogenic species that drive neuronal dysfunction rather the large amyloid aggregates. Oligomeric soluble forms of Abeta have toxic properties and are believed to induce synaptic degeneration. Active zones are the sites along nerve terminals where synaptic vesicles dock and undergo exocytosis during synaptic transmission. They are characterized by active endocytosis and exocytosis. There are studies suggesting that cellular membranes play a major role in Abeta oligomerization process. However, interaction of Abeta peptides with the plasma membrane, endocytosis of Abeta oligomers and the effect on membrane dynamics at the synapse level is still poorly understood. Activity report of months 1-12 (scientist in charge: Dr. ML Maddelein) We investigated the link between in vitro amyloid structure and the capability of Abeta peptides to be internalized in neuronal cells. By confocal imaging and Congo red staining, we analyzed the binding at the plasma membrane, internalization and further elongation of oligomers Abeta wild type oligomers (Ab42) compared to a modified Abeta peptide (Ab10Para - affected in its elongation step). Contrary to what was obtained for Ab42, the majority of the mutant Ab10Para was internalized. Moreover, we observed that small oligomers further elongate, not only at the plasma membrane but also in endocytic compartments producing oligomers of specific size inside the cells. Inhibition of Abeta aggregation either by antibody or by inhibitor binding, affects both adhesion at the plasma membrane and cell toxicity. In addition, the modified Ab10Para -unable to elongate- showed a low level of stress induced toxicity. Activity report of months 13-18 (scientist in charge Dr. M Khrestchatisky) We further characterized the aggregation properties of the mutant Ab10Para in vitro and in cellulo. In summary, our results underline the elements involved in Ab aggregation, such as GM1. We found that Ab10Para acts a dominant negative in amyloidogenesis by competiing for the binding at the cell surface, blocking the formation of toxic oligomers, and inhibiting ROS formation. These results are the subject of a manuscript in preparation. These studies open the way for the understanding of Ab-membrane interaction in Ab-induced neurotoxicity and for the design of peptides aimed to inhibit Ab aggregation and neurotoxicity. We analyzed the effect of Ab aggregation on the endocytic trafficking of RAGE. We found that Ab42 induces the formation of clusters of RAGE at the cell surface. These results confirm our hypothesis that RAGE participates in Ab42 aggregation and are important to elucidate the molecular basis of Ab propagation. We are currently analyzing the effect of Ab42 FAD mutants and of Ab10Para on RAGE endocytic trafficking. We expect that the various mutant differentially affects RAGE endocytic trafficking. We demonstrated that RAGE activation leads to TXNIP expression, which is necessary to mediate RAGE function. We found that Ab42 induces the expression of TXNIP in endothelial and neuronal cells, as well as in astrocytes. Moreover, we have preliminary results indicating that TXNIP is over-expressed in the hippocampus of a transgenic mice model of Alzheimer's disease (AD). We produced the tools to inhibit the expression of TXNIP in various cell types and in vivo. The characterization of these tools resulted in 2 peer reviewed publications. These tools are necessary to validate the role of TXNIP in AD progression and we are currently investigating the function of TXNIP in AD progression using these tools. These studies will be very important to understand the molecular mechanisms responsible for Ab-induced toxicity.