Objective: To characterize microglial function and inflammatory mechanisms in LRRK2-associated Parkinson’s disease (PD).

Background: PD is a neurodegenerative disease affecting primarily dopaminergic neurons of the substantia nigra pars compacta. Mutations in LRRK2 cause the most frequent genetic form of PD. While neuronal phenotypes are commonly being investigated in the context of LRRK2-PD, inflammatory processes and the activation of microglia have also been described in different disease models.

Method: We used an established differentiation protocol to derive microglia from iPSCs of affected and unaffected LRRK2 G2019S mutation carriers as well as healthy and isogenic controls. To trigger an inflammatory response, the cultures were exposed to the cytokine interferon gamma (IFN-γ). Before and after treatment, we assessed the LRRK2 phosphorylation status by means of Western blotting, performed immunocytochemistry of PD-specific targets, and subjected the cultures to RNA sequencing to evaluate differentially expressed genes and pathways.

Results: LRRK2 G2019S microglia display higher levels of phosphorylation at serine 1292 (S1292) upon IFN-γ treatment compared to healthy control and gene-corrected cells, suggestive of increased LRRK2 kinase activity upon inflammatory stimuli in mutant cells. We were able to rescue this phenotype by treating G2019S-mutant microglia with MLi-2 – a potent LRRK2 kinase inhibitor. In addition, we found significantly higher levels of α-synuclein in microglia derived from manifesting PD patients after stimulation with IFN-γ compared to, both, microglia from non-manifesting G2019S mutation carriers and gene-corrected control cells, providing insights into mechanisms of disease manifestation. Finally, our RNAseq dataset revealed proinflammatory signaling in LRRK2-mutant microglia.

Conclusion: Upon inflammatory stimulus, S1292-LRRK2 is a mutation-specific marker of LRRK2 kinase activity, while endogenous levels of microglial α-synuclein pose a possible mechanism of disease manifestation in our iPSC-derived microglia models. Our data highlights microglia as significant contributors to the pathogenesis of LRRK2-PD.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/investigating-inflammatory-mechanisms-in-ipsc-derived-lrrk2-microglia/