Developing iPSC-derived neuronal cell lines for studying the effect of synuclein overexpression in the context of synucleinopathies

Abstract

Synucleinopathies are a group of neurodegenerative disorders that share a common pathological lesion composed of intracellular aggregates of α-synuclein (α-syn). This study focuses on two diseases in this group: Parkinson’s disease (PD) and dementia with Lewy bodies (DLB). Both pathologies are characterized by α-syn aggregates in the neurons, however they have distinct characteristics with respect to the cell type containing the aggregates and the predominantly affected brain region. Specifically, the dopaminergic neurons are primarily affected in PD whereas DLB is largely defined by aggregates in the cholinergic neurons. α-syn gene (SNCA) has been implicated in the etiology of this group of diseases and accumulating evidence has suggested that overexpression of SNCA plays an important role in the disease development. Our goal is to develop a biological model system using induced Pluripotent Stem Cells (iPSCs) to compare the effect of SNCA overexpression among the pathologies of PD and DLB. The first step towards the accomplishment of this goal was to establish a protocol to differentiate iPSCs into dopaminergic neurons (mDA) and Basal Forebrain Cholinergic Neurons (BFCN). iPSCs from a patient caring the genomic triplication of SNCA locus and iPSCs from a control (normal Karyotype) were used for the development of this model system. iPSCs were successfully differentiated into mDAs and BFCNs and neurons were characterized using RT-PCR, quantitative real-time PCR, and immunofluorescence. This prototype was useful to functionally characterize the effects of SNCA triplication on mDAs and BFCNs, on both molecular and cellular levels. Taking this model into future research, we will attempt to use these models to elucidate the functional effects of SNCA overexpression on specific neuronal types involved in the different synucleinopathies.