Characterisation of oxidative stress, dna damage and inflammation in a cellular model of parkinson's disease

Abstract

Background: Parkinson's disease (PD) is the second most common neurodegenerative disease and is a synucleinopathy due to the critical role of α-synuclein (α-Syn) in its pathology. α-Syn is able to translocate from the cytoplasm to the nucleus and cause DNA damage. Methods: SH-SY5Y cells were stably transfected with plasmids containing wild type (WT) α-Syn and A53T mutant α-Syn as fusion proteins with EGFP and an EGFP only control vector. The cells were differentiated using retinoid acid (RA) and treated with hydrogen peroxide (H2O2) and analysed in a differentiated state for the effects of oxidative stress using flow cytometry, DNA damage using γH2A.X staining and inflammatory and senescence markers using qPCR. Results: Cells expressing the A53T mutation exhibited a higher sensitivity to treatment with H2O2 with significantly higher amounts of DNA damage. Intriguingly, WT α-Syn seemed to have a protective effect in differentiated cells under increased stress with no increase in DNA damage after H2O2 treatment. In line with these results, inflammatory markers of COX- 2, IL-6 and TNF-α as well as the stress-and senescence marker p21 were all significantly increased in differentiated treated cells expressing mutant α-Syn compared to cells harbouring WT α-Syn or EGFP controls only. Interestingly, both forms of α-Syn equally increased the levels of mitochondrial ROS already in untreated condition. Conclusions: Our data demonstrate a higher stress sensitivity of SH-SY5Y neuroblastoma cells harbouring an A53T mutant α-Syn for DNA damage and the expression of selected inflammatory marker. Intriguingly, WT α-Syn seemed to provide a protection against oxidative stress in our model which was most pronounced for the absence of increased DNA damage after hydrogen peroxide treatment. Although the mechanism of protection is not clear yet, we suggest that the prevention of DNA damage correlates to less neuroinflammation, an area of intense research in PD pathology.