Objective: The aim of the present study was to assess biochemical markers in primary macrophages that could distinguish patient with Parkinson’s disease linked to mutations in the glucocerebrosidase (GBA) gene (GBA-PD) from healthy carriers of GBA mutations (GBA-carriers).

Background: Not all GBA mutation carriers develop PD during their lives. Search for PD biomarkers in GBA mutation carriers remains important. Macrophages exhibit the highest level of glucocerebrosidase (GCase) activity and are the most vulnerable for GCase deficiency.

Method: GCase activity, hexosylsphingosine (HexSph), GCase level, level of GCase translocation to lysosomes and degree of autophagy were assessed in primary macrophages from GBA-PD (N=14), GBA-carriers (N=14) and controls (N=32). Peripheral blood mononuclear cells-derived primary macrophages were cultured as described earlier [1]. GCase enzymatic activity and HexSph concentration were measured by liquid chromatography with tandem-mass spectrometry (LC-MS/MS) in dry macrophage cell spots. GCase translocation (Alexa Fluor 488) to lysosomes (marker LAMP2 (Cy3)) was assessed using a Leica TCS-SP5 confocal microscope. Primary anti-GBA and anti-GAPDH antibodies were used to determine relative GCase level [1]. Alpha-synuclein level was determined by ELISA using Human alpha-synuclein ELISA kit (Thermo Fisher Scientific, USA) [2]. Spontaneous autophagy was evaluated using Autophagy Detection kit (Abcam, UK).

Results: All measured parameters were altered as in GBA-PD patients so in GBA-carriers compared to controls independent of the disease status [Table 1]. A decrease in GCase activity, an increase in HexSph concentration, a decrease in GCase level and GCase translocation to lysosome, an increase of alpha-synuclein level and autophagy disturbances were shown GBA-carriers compared to controls. The only difference between GBA-PD patients and GBA-carriers was in HexSph concentration (p<0.05)

Conclusion: GBA mutations lead to a decrease in GCase activity, GCase level and an efficiency of GCase translocation to lysosome, an increase of alpha-synuclein level and autophagy disturbances in primary macrophages independent from PD status. HexSph concentrations could be viewed as a possible biochemical marker of PD in GBA mutations carriers. The study was funded by RSF №19-15-00315.

References: 1.Kopytova AE, Rychkov GN, Nikolaev MA, Baydakova GV, Cheblokov AA, Senkevich KA, Bogdanova DA, Bolshakova OI, Miliukhina IV, Bezrukikh VA, Salogub GN, Sarantseva SV, Usenko TC, Zakharova EY, Emelyanov AK, Pchelina SN. Ambroxol increases glucocerebrosidase (GCase) activity and restores GCase translocation in primary patient-derived macrophages in Gaucher disease and Parkinsonism. Parkinsonism Relat Disord. 2021 Mar;84:112-121. doi: 10.1016/j.parkreldis.2021.02.003. Epub 2021 Feb 10. PMID: 33609962.
2. Emelyanov A, Usenko T, Nikolaev M, Senkevich K, Kulabukhova D, Lavrinova A, Andoskin P, Miliukhina I, Pchelina S. Increased α-Synuclein Level in CD45+ Blood Cells in Asymptomatic Carriers of GBA Mutations. Mov Disord. 2021 Aug;36(8):1997-1998. doi: 10.1002/mds.28688. PMID: 34409693.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/biochemical-markers-of-glucocerebrosidase-gcase-deficiency-in-primary-macrophages-derived-from-gba1-mutation-carriers-with-and-without-parkinsons-disease/