Abstract
Understanding the specific gene changes underlying the prodromic stages of Alzheimer’s disease pathogenesis will aid the development of new, targeted therapeutic strategies for this neurodegenerative disorder. Here, we employed RNA-sequencing to analyze global differential gene expression in a defined model nerve cell line expressing α4β2 nicotinic receptors (nAChRs), high-affinity targets for beta amyloid (Aβ). The nAChR-expressing neuronal cells were treated with nanomolar Aβ1–42 to gain insights into the molecular mechanisms underlying Aβ-induced neurotoxicity in the presence of this sensitizing target receptor. We identified 15 genes (out of 15,336) that were differentially expressed upon receptor-linked Aβ treatment. Genes up-regulated with Aβ treatment were associated with calcium signaling and axonal vesicle transport (including the α4 nAChR subunit, the calcineurin regulator RCAN3, and KIF1C of the kinesin family). Downregulated genes were associated with metabolic, apoptotic or DNA repair pathways (including APBA3, PARP1 and RAB11). Validation of the differential expression was performed via qRT-PCR and immunoblot analysis in the defined model nerve cell line and primary mouse neurons. Further verification was performed using immunocytochemistry. In conclusion, we identified apparent changes in gene expression on Aβ treatment in the presence of the sensitizing nAChRs, linked to early-stage Aβ-induced neurotoxicity, which may represent novel therapeutic targets.
Highlights
Amyloid-β (Aβ) is a short, potentially neurotoxic peptide derived from amyloid precursor protein (APP) in select regions of the brain[1,2]
Regulated genes were examined in Aβ-treated mouse hippocampal neurons as a validating primary in vitro neuronal model endogenously expressing nAChRs and in 5xFAD APP/presenilin 1 (PS1) mutant mouse hippocampus
As a defined in vitro neuronal model expressing one of the prominent receptor targets for Aβ, namely high affinity α4β2-type nicotinic receptors, which sensitize the cells to Aβ toxicity[14], neuroblastoma hybrid rodent NG108-15 cells exclusively expressing mouse α4β2-nAChRs were treated daily with 100 nM soluble oligomeric Aβ1–42 as compared to vehicle-treated, receptor-expressing controls
Summary
Amyloid-β (Aβ) is a short, potentially neurotoxic peptide derived from amyloid precursor protein (APP) in select regions of the brain[1,2]. While changes in Aβ levels are only correlative with stages of AD, understanding differential gene expression related to Aβ-induced toxicity pathways upon Aβ binding to known target receptors may provide new tools for study focused on Aβ neurotoxicity. We further demonstrated that this nAChR-induced Aβ neurotoxicity occurs through the timed alteration of discrete intracellular signaling molecules[14]. This prompted our study to investigate differential changes in downstream pathways underlying Aβ-linked neurotoxicity at a genetic level, possibly revealing new cellular targets for intervention in neurodegenerative processes. Regulated genes were examined in Aβ-treated mouse hippocampal neurons as a validating primary in vitro neuronal model endogenously expressing nAChRs and in 5xFAD (familial Alzheimer’s disease) APP/presenilin 1 (PS1) mutant mouse hippocampus
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