Objective: To characterize neuroinflammatory gene expression in microglia and astrocytes associated with early alpha-synuclein (a-syn) inclusion formation.

Background: Parkinson’s disease (PD) is characterized by alpha-synuclein (a-syn) inclusions, the progressive loss of dopamine neurons in the substantia nigra pars compacta (SNpc) and neuroinflammation. These pathological features can be recapitulated in vivo using the alpha-synuclein preformed fibril (PFF) model. Specifically, the accumulation of phosphorylated a-syn (pSyn) inclusions in the SNpc peaks at 2 months following intrastriatal PFF injection, accompanied by a similar peak in reactive microglia and astrocytes, with SNpc degeneration observed at 5-6 months. Using RNA-Seq to analyze transcript levels in the PFF seeded SNpc at 2 months, we observe upregulation of multiple transcripts associated with innate and adaptive immune response pathways. In the present study we establish the reproducibility of these neuroinflammatory gene expression changes and identify the cellular source (microglia vs. astrocytes).

Method: Droplet digital PCR (ddPCR) or fluorescent in situ hybridization (FISH) was used to examine gene expression of individual neuroinflammatory genes in rat nigral tissue at 2, 4 or 6 months after intrastriatal PFF injection.

Results: At the peak of nigral pSyn inclusions we observed increased expression of genes associated with reactive microglia (Stat1, Axl, Casp1, Il18), A1 astrocytes (C3, GBP2, Serping1) and the adaptive immune system (Cd4) using ddPCR.  FISH revealed that microglia in close proximity to SNpc neurons with pSyn inclusions express CD74, C3, and C1qa, whereas Serping1 was expressed by astrocytes. Since C3 expression in the PFF model has previously been attributed to A1 astrocytes, we analyzed C3 expression at 4 and 6 months. Microglia expressed C3 at the 2 and 4 month time point however, expression of C3 switched to astrocytes at 6 months, following degeneration of SNpc neurons.

Conclusion: Our results identify a specific, reproducible phenotype of microglia and astrocytes in association with pSyn inclusions in the SNpc and across the progression of degeneration. Understanding the specific neuroinflammatory response of microglia and astrocytes in the context of a-syn inclusion triggered degeneration may uncover novel therapeutic targets.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/inflammatory-microglial-and-astroglial-phenotype-associated-with-pathological-alpha-synuclein-inclusions-in-the-substantia-nigra/