The role of amyloid beta 4-42 in the etiology of Alzheimer's disease Doctoral Thesis In partial fulfillment of the requirements for the degree “Doctor rerum naturalium (Dr. rer. nat.)” in the Molecular Medicine Study Program at the Georg-August University Göttingen submitted by Yvonne Bouter born in Peine, Germany Göttingen 2014 Members of the thesis commitee: Prof. Dr. Thomas A. Bayer (Supervisor) Department for Psychiatry Division of Molecular Psychiatry University Medical Center Göttingen Göttingen Prof. Dr. Uwe-Karsten Hanisch Dept. of Neuropathology University Medical Center Göttingen Göttingen Prof. Dr. Dr. Hannelore Ehrenreich Division of Clinical Neurosciences Max Planck Institute of Experimental Medicine Göttingen Date of the oral examination: 12.11.2014 Declaration: Here I declare that my doctoral thesis entitled "The role of amyloid beta 4-42 in the etiology of Alzheimer's disease" has been written independently with no other sources and aids than quoted. Yvonne Bouter Göttingen, September 2014 List of Publications: Parts of this thesis have been published: Original articles: Bouter, Y., Dietrich, K., Wittnam, J.L., Rezaei-Ghaleh, N., Pillot, T., Papot-Couturier, S., Lefebvre, T., Sprenger, F., Wirths, O., Zweckstetter, M., and Bayer T.A. (2013). N- truncated amyloid β (Aβ) 4-42 forms stable aggregates and induces acute and long- lasting behavioral deficits. Acta Neuropathol. 126, 189-205. Bouter, Y., Kacprowski, T., Weissmann, R., Dietrich, K., Borgers, H., Brauß, A., Sperling, C., Wirths, O., Albrecht, M., Jensen, L.R., Kuss, A.W., and Bayer T.A. (2014). Deciphering the Molecular Profile of Plaques, Memory Decline and Neuron Loss in Two Mouse Models for Alzheimer's Disease by Deep Sequencing. Front. Aging Neurosci. 6. Antonios, G., Saiepour, N., Bouter, Y., Richard, B.C., Paetau, A., Verkkoniemi-Ahola, A., Lannfelt, L., Ingelsson, M., Kovacs, G.G., Pillot, T., Wirths, O., and Bayer T.A. (2013). N- truncated Abeta starting with position four: early intraneuronal accumulation and rescue of toxicity using NT4X-167, a novel monoclonal antibody. Acta Neuropathol Commun 1, 56. Guzmán, E.A., Bouter, Y., Richard, B.C., Lannfelt, L., Ingelsson, M., Paetau, A., Verkkoniemi-Ahola, A., Wirths, O., and Bayer T.A. (2014). Abundance of Aβ5-x like immunoreactivity in transgenic 5XFAD, APP/PS1KI and 3xTG mice, sporadic and familial Alzheimer’s disease. Mol Neurodegener 9, 13. Abstracts: Bouter, Y., Dietrich, K., Wittnam, J.L., Pillot, T., Papot-Couturier, S., Lefebvre, T., Sprenger, F., Wirths, O., Zweckstetter, M., and Bayer T.A. (2013). Tg4-42: A new mouse model of Alzheimer's disease - N-truncated amyloid β (Aβ) 4-42 induces severe neuron loss and behavioral deficits. Alzheimer's and Dementia 9, P498. Dietrich, K., Bouter, Y., Wittnam, J.L., Pillot, T., Papot-Couturier, S., Lefebvre, T., Sprenger, F., Wirths, O., Janc O.A., Müller M., Zweckstetter, M., and Bayer T.A. (2013). Tg4-42: A new mouse model of Alzheimer's disease—N-truncated beta-amyloid 4-42 affects memory decline and synaptic plasticity. Alzheimer's and Dementia 9, P498. Bayer T.A., Bouter Y., Kacprowski T, Sperling C, Albrecht M, Weißmann R , et al. (2014). Deep sequencing unravels the molecular signatures in two different mouse models for Alzheimer disease. Medizinische Genetik. 26, 107. Contents ACKNOWLEDGEMENTS ...................................................................................................... I ABSTRACT ........................................................................................................................ II LIST OF FIGURES .............................................................................................................. IV LIST OF TABLES ................................................................................................................ VI LIST OF ABBREVATIONS .................................................................................................. VII 1 INTRODUCTION ................................................................................................................ 1 1.1 ALZHEIMER'S DISEASE .............................................................................................................. 1 1.2 CLINICAL ASPECTS OF ALZHEIMER'S DISEASE ................................................................................ 1 1.2.1 Epidemiology .................................................................................................................. 1 1.2.2 Non-genetic risk factors ................................................................................................. 1 1.2.3 Disease progression ....................................................................................................... 2 1.2.4 Diagnosis and treatment ................................................................................................ 3 1.2.4.1 Diagnosis ................................................................................................................ 3 1.2.4.2 Treatment ............................................................................................................... 3 1.3 NEUROPATHOLOGICAL HALLMARKS OF ALZHEIMER'S DISEASE ......................................................... 4 1.3.1 Amyloid plaques ............................................................................................................. 4 1.3.2 Neurofibrillary Tangles ................................................................................................... 5 1.3.3 Inflammation .................................................................................................................. 6 1.3.4 Neuron loss..................................................................................................................... 7 1.4 THE AMYLOID PRECURSOR PROTEIN............................................................................................ 7 1.4.1 APP processing ............................................................................................................... 8 1.4.1.1 Non-amyloidogenic pathway ................................................................................. 8 1.4.1.2 Amyloidogenic pathway ....................................................................................... 10 1.5 THE AMYLOID CASCADE HYPOTHESIS......................................................................................... 10 1.5.1 Intraneuronal amyloid hypothesis ............................................................................... 11 1.6 AMYLOID BETA AGGREGATION ................................................................................................ 14 1.7 AMYLOID BETA VARIANTS ....................................................................................................... 15 1.7.1 Truncated amyloid beta species................................................................................... 15 1.7.1.1 Amyloid beta 4-42 ................................................................................................ 17 1.7.2 Modified amyloid beta species .................................................................................... 17 1.8 TRANSGENIC MOUSE MODELS OF ALZHEIMER'S DISEASE .............................................................. 19 1.8.1 The 5XFAD mouse model ............................................................................................. 21 1.9 GENETICS ............................................................................................................................ 22 1.9.1 From genome-wide association studies to next-generation sequencing .................... 24 1.9.2 Transcriptome profiling: RNA-Sequencing ................................................................... 26 1.10 PROJECT OBJECTIVES ............................................................................................................. 27 2 MATERIAL AND METHODS .............................................................................................. 30 2.1 ANIMALS ............................................................................................................................. 30 2.1.1 General considerations and housing ............................................................................ 30 2.1.2 Tg4-42 transgenic mice ................................................................................................ 30 2.1.3 5XFAD transgenic mice ................................................................................................. 31 2.1.4 Preparation of mouse brain tissue for biochemistry ................................................... 32 2.1.5 Preparation of mouse brain tissue for immunohistochemistry ................................... 32 2.1.5.1 Drop-fixation ........................................................................................................ 32 2.1.5.2 Perfusion .............................................................................................................. 32 2.2 BEHAVIOR ANALYSES OF MICE ................................................................................................. 33 2.2.1 Paw-clasping test .......................................................................................................... 34 2.2.2 String suspension ......................................................................................................... 34 2.2.3 Balance Beam ............................................................................................................... 34 2.2.4 Cross maze ..................................................................................................................