Apolipoprotein E and Alzheimer's diseases : Signals and effects

Sammanfattning: Alzheimer's Disease (AD) is the most frequent cause of dementia in the elderly. A diagnosis of AD is confirmed post-mortem by the accumulation of neuritic plaques (NP) and neurofibrillary tangles (NFTs) in brain tissue. NPs are extracellular deposits composed mainly of amyloid beta-peptide (A-beta) that is derived from the P amyloid precursor protein (APP). NFTs are found inside neurons and are characterised by structures referred to as paired helical filaments, whose major component is an abnormally phosphorylated and glycosylated form of the 'tau' protein. The most important risk factor for AD is polymorphism in the apolipoprotein E (apoE) gene. The apoE gene polymorphism gives rise to three different protein isoforms that differ in one single amino acid substitution and which show different lipid binding properties and three dimensional structure. The dose of the apoE4 isoform both increases the risk and reduces the age of onset for AD. The molecular mechanisms underlying this association are not fully understood, and in many cases are controversial. The present thesis, focuses on investigating several aspects of apoE biology in the context of AD. Paper I characterised factors that regulate apoE secretion in brain. ApoE secretion was shown to be modulated by adenosine 3':5'-cyclic monophosphate (cAMP) and factors that regulate intracellular cAMP. Retinoic acid potentiated, while the protein kinase C (PKC) activator phorbol 12- myristate 13-acetate (PMA) reversed the cAMP mediated effect. High PMA concentrations decreased apoE secretion. Also several agonists linked to the phospholipase C/PKC signalling pathway had opposite effects on apoE secretion, suggesting that the regulation of apoE by PKC may result from both positive and negative mechanisms. PKC activation and cAMP had opposite effects on nerve growth factor secretion than those seen for apoE secretion. Paper II aimed to clarify the interactions between A-beta, apoE isoforms and apoE containing lipoproteins with respect to neurotoxicity. In human SH-SY5Y neuroblastoma cells, both apoE3 and E4 were toxic when used alone but not when associated with P-very low density lipoproteins (beta-VLDL). Beta-VLDL also reduced the toxicity of A beta. These results suggest that the conformation of lipoprotein containing apoE particles may be important for determining the direct apoE contribution to neurotoxicity. No toxic effects were found in human fibroblasts, suggesting cell specificity. Papers III, IV and V explored the hypothesis that apoE may act as modulator of intracellular signalling pathways implicated in AD pathology. Paper III showed that apoE activates PKC without either isoform specific effects or further consequences for alpha-secretase APP cleavage. Paper IV showed that apoE4 and A-beta(1-42), but not apoE3, disrupt carbachol-induced phosphoinositide (P1) hydrolysis by an oxidative stress mediated mechanism. Moreover, apoE3 protected against the A-beta effect. Both apoE4 and A-beta(1-42) mediated disruptions of PI hydrolysis were protected by estrogen via an antioxidant effect and activation of phosphatidylinositol 3-kinase (PI3K). These findings may help to explain the lower effectiveness of treatments for AD based on cholinergic enhancers in apoE4 carriers, as well as shedding light on the protective mechanism of action of estrogen in AD. Paper V showed apoE isoform differences on the regulation of glycogen synthase kinase-3 beta (GSK-3 beta) activity. ApoE3 induced a mild transient early activation of GSK-3 beta that was possibly balanced by PKC activation. In contrast, apoE4 regulated GSK-30 activity in a biphasic manner, with a strong early activation and a subsequent inactivation of GSK-3 beta. ApoE4 also activated PKC-alpha and PKB, which may have given the subsequent GSK-3 beta inhibition. A beta(1-42) modulation of GSK-3 beta activity was also biphasic with a strong early activation and a subsequent inactivation. A beta(1-42) also induced an early and potent activation of PKC-alpha and a decreased PKB activity. These studies provide insight into some of the factors that control apoE secretion in brain and the neurotoxic effects of variant apoE isoforms. They also provide evidence that apoE has isoform specific effects on several signalling cascades involved in A beta production, tau phosphorylation, and apoptosis.