Objective: To assess glucocerebrosidase and alpha-synuclein level in primary macrophages derived from patients with Parkinson’s disease with and without GBA1 mutation.
Background: Parkinson’s disease (PD) is a neurodegenerative disorder that is characterized by the accumulation of abnormal protein aggregates of alpha-synuclein in the brain. Mutations in the GBA1 gene, encoding the lysosomal enzyme glucocerebrosidase (GCase), are the most common genetic risk factor for PD. GCase is the key enzyme in ceramide metabolism, the enzyme hyderolyzes glucose moieties from glucosylceramide and glucosylsphingosine. Some studies demonstrated that reduced GCase activity might be resulted in increased alpha-synuclein protein. Monocyte-derived macrophages represent one of the most promising models for investigating molecular mechanisms of GCase dysfunction, as this cell type is vulnerable for disturbances in ceramide metabolism.
Method: Mononuclear fraction was isolated from whole blood of patients with GBA1 mutation with (GBA1-PD) (N=10), sporadic PD (sPD) (N=12) and controls (N=25). Primary macrophages were cultured using RPMI-1640 supplemented with 10% FBS, 1% gentamicin and 10 ng/ml M-CSF for 4 days, with daily media changes. Total protein (10μg) was separated by SDS-PAGE and then transferred to PVDF membranes by electroblotting. Primary anti-GBA and anti-GAPDH antibodies were used. Digital images were obtained by the chemiluminescence system ChemiDoc. Alpha-synuclein level was determined by ELISA using Human alpha-synuclein ELISA kit (Thermo Fisher Scientific, USA).
Results: In present study we showed an increased alpha-synuclein level in primary macrophages from GBA1-PD 1,14 (0,27 – 5,99) ng/µg and sPD patients 2,34 (0,37 – 4,61) ng/µg compared to controls 0,41 (0,09 – 1,60) (p=0.005, p<0.001, corresponding). Moreover, a decreased relative GCase protein level was observed in sPD primary macrophages 0,78 (-0,09 – 1,85) compared to controls 1,82 (0,46–3,74) (p=0.025).
Conclusion: Patients with Parkinson’s disease are characterized by increased alpha-synuclein protein level in primary macrophages regardless of the status of the GBA1 mutations. It is interesting to note that decreased relative GCase level in sPD patients may be triggered by the accumulation of alpha-synuclein.
The study was funded by RSF №19-15-00315.
To cite this abstract in AMA style:
A. Kopytova, M. Nikolaev, G. Baydakova, A. Izymchenko, D. Bogdanova, I. Miliukhina, A. Emelyanov, E. Zakharova, S. Pchelina. Increased alpha-synuclein level in primary macrophages derived from patients with Parkinson’s disease [abstract]. Mov Disord. 2022; 37 (suppl 2). https://www.mdsabstracts.org/abstract/increased-alpha-synuclein-level-in-primary-macrophages-derived-from-patients-with-parkinsons-disease/. Accessed September 7, 2024.« Back to 2022 International Congress
MDS Abstracts - https://www.mdsabstracts.org/abstract/increased-alpha-synuclein-level-in-primary-macrophages-derived-from-patients-with-parkinsons-disease/