Caspase Activation in an In-Vitro Model of Parkinson’s Disease

Parkinson’s disease (PD) is the second most prevalent neurodegenerative disorder worldwide with a key hallmark of its pathology being the degeneration of dopaminergic neurons in the substantia nigra (SNc) due to intraneuronal accumulation of Lewy bodies. Lewy bodies form through the aggregation of the monomer alpha-synuclein protein (aSyn), which can misfold into toxic fibrils that disrupt cellular homeostasis and cause apoptosis. While there is no cure for PD, establishing good in-vitro models is essential to further research of the disease and to developing possible treatments and therapies. This study’s goal was to develop an in-vitro model of PD using SH-SY5Y cells that would allow us to test the properties of our DNA-aptamer, a-syn-1, in preventing aSyn aggregation and development of pathology. This model was created by differentiating the SH-SY5Y cells into N-type neuroblast-like cells using retinoic acid following either Protocol A (full media with 10% FBS) or Protocol B (reduced media with 1% FBS) before adding aSyn filbrils. The cells were then stained with a Caspase-3/7 Green reagent and a NucBlue counterstain and were imaged at 24 h, 48 h, 72 h, and 96 h time points. We found that Protocol A showed no significant caspase activation at any of the timepoints and became overgrown with S-type epithelial-like cells after 94 h. Protocol B showed less cell proliferation and less S-type cells at the 94 h time point. Future studies will perform caspase testing on Protocol B to determine if it is the ideal in-vitro PD model before adding a-syn-1.