Derivation, Characterization, and Neural Differentiation of Integration-Free Induced Pluripotent Stem Cell Lines from Parkinson’s Disease Patients Carrying SNCA, LRRK2, PARK2, and GBA Mutations

Patrick Lewis,Olga Momcilovic,Mahendra S Rao,Morten Meyer,Xianmin Zeng,Renuka Sivapatham,Sean Mooney,Tal Ronnen Oron

doi:10.1371/journal.pone.0154890

Patrick Lewis, Olga Momcilovic + Show 6 more

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https://doi.org/10.1371/journal.pone.0154890

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Abstract

We report generation of induced pluripotent stem cell (iPSC) lines from ten Parkinson’s disease (PD) patients carrying SNCA, PARK2, LRRK2, and GBA mutations, and one age-matched control. After validation of pluripotency, long-term genome stability, and integration-free reprogramming, eight of these lines (one of each SNCA, LRRK2 and GBA, four PARK2 lines, and the control) were differentiated into neural stem cells (NSC) and subsequently to dopaminergic cultures. We did not observe significant differences in the timeline of neural induction and NSC derivation between the patient and control line, nor amongst the patient lines, although we report considerable variability in the efficiency of dopaminergic differentiation among patient lines. We performed whole genome expression analyses of the lines at each stage of differentiation (fibroblast, iPSC, NSC, and dopaminergic culture) in an attempt to identify alterations by large-scale evaluation. While gene expression profiling clearly distinguished cells at different stages of differentiation, no mutation-specific clustering or difference was observed, though consistent changes in patient lines were detected in genes associated mitochondrial biology. We further examined gene expression in a stress model (MPTP-induced dopaminergic neuronal death) using two clones from the SNCA triplication line, and detected changes in genes associated with mitophagy. Our data suggested that even a well-characterized line of a monogenic disease may not be sufficient to determine the cause or mechanism of the disease, and highlights the need to use more focused strategies for large-scale data analysis.

Highlights

Parkinson's disease (PD) is a chronic, progressive and devastating neurodegenerative disorder, characterized by a profound loss of nigrostriatal dopaminergic neurons and accumulation of misfolded α-synuclein protein aggregates called Lewy bodies in remaining dopaminergic neurons and in other brain areas such as the cortex [1]
We describe the derivation, characterization, and neural differentiation of eight integration-free induced pluripotent stem cell (iPSC) lines derived from seven PD patients carrying various mutations, including SNCA, LRRK2, PARK2, and GBA, and an age-matched control
Given the importance of SNCA in PD etiology, we examined gene expression in the SNCA line followed by MPTP treatment and found significant changes in genes associated to mitochondrial biology and cell death

Summary

Introduction

Parkinson's disease (PD) is a chronic, progressive and devastating neurodegenerative disorder, characterized by a profound loss of nigrostriatal dopaminergic neurons and accumulation of misfolded α-synuclein protein aggregates called Lewy bodies in remaining dopaminergic neurons and in other brain areas such as the cortex [1]. Multiple genes, including PARK2, LRRK2, GBA, SNCA, DJ-1, and PINK-1, have been linked to familial forms of PD (for review see [3,4,5,6]). The most common monogenic PD-associated mutation is a G2019S substitution in LRRK2 that causes the neurotoxic gain of function of LRRK2 protein kinase [7,8]. Many autosomal recessive mutations in PARK2 have been detected and account for the most early onset PD cases [5]. Mutations in GBA that are the causative factor for Gaucher disease, a lysosomal storage disease, are associated with Lewy body pathology and PD [9]

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