Parkinson’s Disease-Associated LRRK2 Interferes with Astrocyte-Mediated Alpha-Synuclein Clearance

Abstract

BackgroundParkinson’s disease (PD) is a neurodegenerative, progressive disease without cure. To prevent PD onset or at least limit neurodegeneration, a better understanding of the underlying cellular and molecular disease mechanisms is crucial. Mutations in the leucine rich repeat kinase 2 ( LRRK2 ) gene represent one of the most common causes of familial PD. In addition, LRRK2 variants are risk factors for sporadic PD, making LRRK2 an attractive therapeutic target. M utations in LRRK2 has been linked to impaired alpha-synuclein (α-syn) degradation in neurons. However, in which way pathogenic LRRK2 affects α-syn clearance by astrocytes, the major glial cell type of the brain, remains unclear. The impact of astrocytes on PD progression has recently received much attention and recent data indicate that astrocytes play a key role in α-syn mediated pathology. MethodsIn the present study, we aimed to compare the capacity of wild-type astrocytes and astrocytes carrying the PD-linked G2019S mutation in Lrrk2 to ingest and degrade fibrillary α-syn. For this purpose, we used two different astrocyte culture systems that were exposed to sonicated α-syn for 24 hours and analyzed directly after the α-syn pulse or 6 days later. To elucidate the impact of LRRK2 on α-syn clearance, we performed various analyses, including complementary imaging, transmission electron microscopy and proteomic approaches. ResultsOur results show that astrocytes carrying the G2019S mutation in Lrrk2 exhibit a decreased capacity to internalize and degrade fibrillar α-syn via the endo-lysosomal pathway. In addition, we demonstrate that the reduction of α-syn internalization in the Lrrk2 G2019S astrocytes is linked to Annexin A2 (AnxA2) loss-of-function. ConclusionTogether, our findings reveal that astrocytic LRRK2 contributes to the clearance of extracellular α-syn aggregates through an AnxA2-dependent mechanism.

Inhibition of Ceramide Synthesis Reduces α-Synuclein Proteinopathy in a Cellular Model of Parkinson’s Disease

“…LPS-free recombinant α-syn was produced and purified as previously described, and α-syn fibrils were obtained by aggregating monomeric α-syn for 7 days at 37 °C with shaking at 1000 rpm (ThermoMixer F1.5 Eppendorf, 1000 rpm) [ 61 ]. α-Syn monomer was resuspended in sterile PBS at 280 µM for the aggregation assay, and after 7 days, fibrils were collected by centrifugation to remove residual soluble monomeric or oligomeric species in the supernatant.…”

Trafficking of the glutamate transporter is impaired in LRRK2-related Parkinson’s disease

“…In this regard, our group recently reported a loss of function of Annexin A2 (AnxA2) in Lrrk2 G2019S astrocytes [75]. AnxA2 is a protein involved in sorting endosome maturation and recycling endosome formation [15,54,75]. Although further studies are warranted, AnxA2 deficits may be at least in part responsible for the enlarged size of the Rab4-positive endosomal recycling structures reported here.…”

“…By combining confocal and TEM ultrastructural analysis, we show that the Lrrk2 G2019S mutation profoundly affects the architecture of the fast-recycling Rab4-positive organelles in astrocytes, promoting an enlargement of the area of these vesicles. In this regard, our group recently reported a loss of function of Annexin A2 (AnxA2) in Lrrk2 G2019S astrocytes [75]. AnxA2 is a protein involved in sorting endosome maturation and recycling endosome formation [15,54,75].…”

“…Mouse primary striatal astrocytes were obtained from postnatal animals between day 1 and day 3 as described in [75]. Brains were removed from their skull and placed in a dish containing cold Dulbecco's Phosphate Buffered Saline (DPBS, Biowest).…”

Extracellular Vesicles as Novel Diagnostic and Prognostic Biomarkers for Parkinson’s Disease

“…Aside the early-onset familial PD, most (≥ 90%) PD cases are sporadic, with current evidence indicating a complex interplay between genetic susceptibility and a panel of environmental factors contributing to PD pathophysiology [ 3 , 12 , 28 - 36 ]. Indeed, several genes - including α-Syn (SNCA), parkin (PRKN), PTEN-induced putative kinase (PINK1), protein deglycase (DJ-1), and leucine-rich-repeat kinase 2 (LRRK2) - and many environmental factors - chiefly aging and inflammation - impact on the regulation of crucial pathways involved in oxidative stress, mitochondrial function, endoplasmic reticulum stress, autophagic catabolism, protein misfolding and aggregation, finally resulting in the progressive loss of DAergic neurons [ 28 , 34 , 37 - 42 ].…”