Neuroprotective Potential of the N-Terminal Beta Amyloid Peptide Fragment in the Neurodegeneration, Synaptic Dysfunction and Memory Deficits in Models of Alzheimer’s Disease.

Alfulaij, Naghum E. R. I.

Neuroprotective Potential of the N-Terminal Beta Amyloid Peptide Fragment in the Neurodegeneration, Synaptic Dysfunction and Memory Deficits in Models of Alzheimer’s Disease.

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2017-12-phd-alfulaij.pdf (9.67 MB)

Date

2017-12

Authors

Alfulaij, Naghum E. R. I.

Department

Cell & Molecular Biology

Abstract

Beta amyloid (Aβ) plays a central role in the pathogenesis of Alzheimer’s disease (AD). It is produced by the sequential cleavage of amyloid precursor protein (APP) by β- and γ-secretases. This amyloidogenic pathway produces peptides 38-42 amino acids in length, based on the sites cleaved by γ-secretase. Aβ1-42 is the predominant peptide species found in neuritic plaques. Its accumulation and impaired clearance are associated with disease progression. An alternative pathway has been proposed wherein short, N-terminal Aβ fragments are produced. The N-terminal fragments are hydrophilic, making them soluble and less likely to aggregate into plaques. Aβ1-15 is the N-terminal Aβ fragment of focus in this study. It is produced by the sequential cleavage of APP by β- and α-secretases followed by a carboxypeptidase. Aβ1-15 is also upregulated in AD patients suggesting a possible compensatory switch between pathways to suppress Aβ1-42 production. We have recently shown that Aβ1-15 can enhance long-term potentiation (LTP) in wild-type hippocampal synapses at very low (fΜ) concentrations. Aβ1-15 was also shown to reverse the LTP block incurred by Aβ1-42, as well as rescue LTP deficits in APPswe mice. This fragment has a potent and effective signaling activity via nicotinic acetylcholine receptors (nAChRs). Taken together, these data support a neuromodulatory function and a possible neuroprotective action for Aβ1-15. This study focuses on the effects of Aβ1-15 on Aβ1-42 toxicity in various neuronal models, including in vitro NG108-15 hybrid neuroblastoma cells, ex vivo hippocampal neuron cultures and in vivo wild-type and AD model-APP mice. We explored Aβ1-15 neuroprotection against Aβ-mediated neurodegeneration in vitro by incubating our differentiated neuroblastoma cell line with different treatment combinations and across different time points in order to examine both the potency of Aβ1-15 as well as the different ways by which Aβ1-15 might be affecting Aβ1-42 toxicity. Specifically, we explored the possibilities of Aβ1-15 priming by pretreating cells with Aβ1- 15 before the addition of the toxic Aβ1-42, competition of the two fragments by combination treatment, or rescue experiments by adding Aβ1-15 after addition of Aβ1-42 to determine whether Aβ1-15 can reverse or halt toxicity caused by Aβ1-42. Cellular toxicity was assessed as oxidative stress (production of reactive oxygen species, ROS) and apoptotic cell death. Next, we investigated the neuroprotective effects of Aβ1-15 ex-vivo by establishing primary hippocampal neuron cultures to confirm neuroprotection in a more physiologically relevant neuronal model. We then explored the potential for the Aβ1-15 to protect or reverse (rescue) synaptic dysfunction and memory deficits resulting from Aβ synaptotoxicity. Changes in synaptic plasticity were assessed by measuring LTP in acute mouse hippocampal slices. Furthermore, we inspected the potential for rescue by Aβ1-15 of LTP in APPswe hippocampal slices known to have LTP deficits. Lastly, we explored the effect of Aβ1-15 in different behavior paradigms on 5XFAD (FAD: familial Alzeimer’s disease) mice, a model expressing APPswe, APP-London and APPFlorida mutant transgenes as well as two mutant presenilin (PS1) transgenes, which accumulates high levels of Aβ over an accelerated timeframe (months). The behaviors examined were those related to deficits observed AD, namely contextual fear conditioning, novel object recognition and elevated plus maze to examine effects on memory processing, recognition memory and anxiety. We were able to show that the Aβ1-15 protected against all measures of Aβ- triggered neurotoxicity and neuronal dysfunction: oxidative stress, DNA fragmentation, apoptotic cell death, synaptotoxicity and behavioral deficits. Notably, Aβ1-15 prevented LTP inhibition caused by Aβ1-42 treatment and, when injected into the hippocampus was able to rescue memory in contextual fear conditioning as well as decrease anxiety in the 5XFAD mice. To address preliminarily the possible molecular mechanisms underlying the rescue by Aβ1-15 of memory deficits in the 5XFAD, we explored signaling pathways known to be involved in Aβ synaptotoxicity. We observed a substantial upregulation of the glutamate receptor GluR2 and phosphorylated CREB in mouse hippocampi injected with the Aβ1-15, giving us insight into the specific actions of N-terminal fragment. In summary, the data show that Aβ1-15 fully protected against Aβ1-42 –induced cellular toxicity, synaptotoxicity and behavioral deficits. Taken together, the data support our hypothesis that the N-terminal fragment (Aβ1-15) is not only neuroprotective against acute Aβ1-42 toxicity, but also has the ability to rescue memory in 5XFAD mice, potentially introducing a new avenue for AD therapeutics.

URI

http://hdl.handle.net/10125/62241

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Ph.D. - Cell and Molecular Biology

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