Objective: The main objectives are 1) the confirmation of exosome isolation, based on iPSC-derived midbrain neuron model, 2) the identification of α-synuclein cargo of PD-patient derived exosomes in comparison with the isogenic control-derived exosomes, 3) identification of differences in exosomal proteins, mRNAs and DNA content between PD patient-derived and control midbrain neurons.

Background: Parkinson disease is the second most frequent neurodegenerative disease, is characterized by impaired dopamine signaling due to the loss of dopaminergic neurons and this neuronal loss is directly linked with accumulation of α-synuclein.
Exosomes are small extracellular vesicles, which are secreted from many cell types and regulate a plethora of biological mechanisms in the recipient cells. Several studies have linked the exosomal function/cargo with PD, as exosomes contain PD-related proteins, mRNAs of PD-related genes and miRNAs, that regulate PD-related spreading

Method: IPSC-derived midbrain neurons of a human carrier of the SNCA Triplication (AST23) and its isogenic control were used as cell model. We set up an efficient exosomal isolation method. Obtained exosomes were characterized with Western blot analysis (WB) and transmission electron microscopy. Finally, we isolated total exosomal RNA and DNA in order to quantify our genes of interest with qPCR and total exosomal extract for WB.

Results: We differentiated AST23 smNPCs and the isogenic control into midbrain neurons, from which we successfully isolated a highly enriched exosomal fraction. Concerning exosomal protein cargo, we confirmed the presence of α -synuclein inside exosomes of both cell lines and we identified differences in the protein levels of lysosomal protease cathepsin D. We identified increased mRNA levels of both PARK7 (DJ1) and PINK1 in the SNCA triplication-derived neuronal exosomes in comparison with the respective isogenic control-derived neuronal exosomes, that were accompanied by increase in the mitochondrial DNA levels.

Conclusion: Our results revealed alterations in the exosomal-sorted lysosomal proteins, PD-related gene transcripts and mtDNA. Yet it is not clear, whether these changes are surrogate biomarkers or may directly related to pathogenic pathway. Further research is needed, as exosomes can reveal new mechanisms of PD spreading, putative PD biomarkers and novel therapeutic targets.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/alterations-in-neuron-derived-exosomal-cargo-in-parkinson-disease/