Cellular and proteomic studies of the mitochondrial ABAD/Aβ complex : investigating its role in Alzheimer's disease

Amyloid beta protein--Analysis;Alzheimer Disease--Pathophysiology;Protein-protein interactions;Cell-mediated cytotoxicity

Taylor, Margaret Alexandra ; Gunn-Moore, Frank J. ; Gunn-Moore, Frank J.

University:University of St Andrews

Department:Biology (School of)

关键词: Amyloid beta protein--Analysis;    Alzheimer Disease--Pathophysiology;    Protein-protein interactions;    Cell-mediated cytotoxicity;   

Others  :  https://research-repository.st-andrews.ac.uk/bitstream/handle/10023/3632/MargaretTaylorPhDThesis.pdf?sequence=3&isAllowed=y

来源: DR-NTU

【 摘 要 】

The focus of this thesis is to investigate the intracellular protein-peptide complex3-hydroxyacyl-CoA dehydrogenase (HADH), also known as ABAD (amyloid-binding alcohol dehydrogenase) and amyloid-beta peptide (Aβ). This complexhas been identified in the development of Alzheimer's disease (AD), and thisstudy tries to identify if ABAD is a useful biomarker for genetic risk profilingstrategies for the early diagnosis of Alzheimer’s disease, or a suitable target fordisease-modifying drug development. The major aim of this project is to explorethe biochemical and cellular processes activated as a result of the interaction ofABAD and Aβ. Understanding the cellular responses to these interactions couldhelp identify important biomarkers and/or drug targets for the diagnosis ortreatment of Alzheimer’s disease. This study assesses the cytotoxic effects of Aβin tissue culture and in animal models overexpressing ABAD.An in vitro cell system using SK-N-SH cells was developed for investigating theeffects of ABAD expression in cells when incubated with synthetic Aβ peptide.In vitro studies confirmed ABAD to be a mitochondrial protein. There wereproblems with the efficiency of the synthetic Aβ peptide used, which was foundto aggregate excessively. Trial of a soluble oligomeric Aβ peptide proved to bemore efficient.A cell system was also developed, culturing neurospheres from murine stem cells.This proved to be a reliable system for culturing primary cells and keeping them in culture for up to 8 weeks. Cells were grown from a wild type strain and thendifferentiated and stained for endogenous expression of proteins.Proteomic studies were carried out with novel transgenic mouse models for AD.Seven proteins were identified with changed expression in the 2x Tg mousemodel. Further immunocytochemistry of human AD brain tissue confirmed theupregulation of peroxiredoxin II and endophilin I. Both proteins could bereturned to normal expression in the mouse models by peritoneal injection for twoweeks of a novel peptide inhibitor to ABAD, confirming the involvement of theABAD/Aβ complex in the increased expression of these proteins.Finally ABAD/Aβ was investigated as a possible target for AD therapy byscreening with a small molecule fragment library. Only an initial screen wascarried out, but several small molecule compounds were found to bind to ABAD.Further screening may produce lead compounds for a synthetic drug to inhibit theenhanced Aβ toxicity associated with the ABAD/Aβ complex.