Abstract
Abstract : iPS reprogramming to model 'disease-in-a-dish has become an attractive approach to study disease mechanisms. However, a frank pathophysiological phenotype in iPSC-derived PD neurons remains to be shown. Objective/Hypothesis: The objective of this proposal is to accelerate the aging process of iPS-derived dopaminergic neurons with the goal of reproducing a Parkinson s disease (PD) specific phenotype in vitro. Specific Aim 1. To generate vector constructs and to establish Dox-inducible iPSC lines that express the Hutchinson-Gilford progeria (HGPS) gene (mutant lamin A with an inframe loss of 150 nucleotides) and differentiate them into dopaminergic neurons. Deliverable of this aim is the introduction of mutant lamin A in iPSC lines and optimization of inducible expression of mutant lamin A in iPSCs and during the differentiation into dopaminergic neurons. Specific Aim 2. To test whether iPSC-derived dopaminergic neurons transduced with mutant lamin A show changes in age-regulated genes at the mRNA and protein levels at different time-points during differentiation, thus exhibiting an accelerated aging. Deliverable of this aim is the assessment of specific age-related aging pattern of gene and protein expression in induced mutant lamin A modified iPSCderived neurons. Specific Aim 3. To test whether induced mutant lamin A cell lines differentiated into dopaminergic neurons exhibit hallmark pathology of PD, such as protein aggregation of alpha-synuclein, posttranslational modification, and signs of mitochondrial pathology. Deliverable of this aim is the assessment of the pathological PD-related phenotype in induced mutant lamin A modified iPSC-derived neurons. Study Design: This is an in vitro study of patient-specific iPSC-derived dopaminergic neurons in which truncated lamin A will be introduced to study cellular phenotypes under the hypothesis that cells under expression of lamin A will age faster than untreated cells.
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