Effects of innate immune receptor stimulation on extracellular \u03b1-synuclein uptake and degradation by brain resident cells

Changyoun Kim,Jerel A Fields,Somin Kwon,Eliezer Masliah,Brian Spencer,Edward Rockenstein,Soo Jean Shin,Michael Mante,Robert A Rissman,Michiyo Iba,Seung-Jae Lee,Anthony Adame

doi:10.1038/s12276-021-00562-6

Abstract

Synucleinopathies are age-related neurological disorders characterized by the progressive deposition of α-synuclein (α-syn) aggregates and include Parkinson’s disease (PD) and dementia with Lewy bodies (DLB). Although cell-to-cell α-syn transmission is thought to play a key role in the spread of α-syn pathology, the detailed mechanism is still unknown. Neuroinflammation is another key pathological feature of synucleinopathies. Previous studies have identified several immune receptors that mediate neuroinflammation in synucleinopathies, such as Toll-like receptor 2 (TLR2). However, the species of α-syn aggregates varies from study to study, and how different α-syn aggregate species interact with innate immune receptors has yet to be addressed. Therefore, we investigated whether innate immune receptors can facilitate the uptake of different species of α-syn aggregates. Here, we examined whether stimulation of TLRs could modulate the cellular uptake and degradation of α-syn fibrils despite a lack of direct interaction. We observed that stimulation of TLR2 in vitro accelerated α-syn fibril uptake in neurons and glia while delaying the degradation of α-syn in neurons and astrocytes. Internalized α-syn was rapidly degraded in microglia regardless of whether TLR2 was stimulated. However, cellular α-syn uptake and degradation kinetics were not altered by TLR4 stimulation. In addition, upregulation of TLR2 expression in a synucleinopathy mouse model increased the density of Lewy-body-like inclusions and induced morphological changes in microglia. Together, these results suggest that cell type-specific modulation of TLR2 may be a multifaceted and promising therapeutic strategy for synucleinopathies; inhibition of neuronal and astroglial TLR2 decreases pathogenic α-syn transmission, but activation of microglial TLR2 enhances microglial extracellular α-syn clearance.

Highlights

Introduction Synucleinopathies such asParkinson’s disease (PD) and dementia with Lewy bodies (DLB) are the second most common age-related neurodegenerative disorders and are characterized by abnormal deposits of α-synuclein (α-syn) called Lewy bodies (LBs) and Lewy neurites (LNs)1–3. αSyn pathology spreads through various brain regions during the progression of the disease[4]
To address the role of innate immune receptors in the kinetics of α-syn fibril uptake, we performed an uptake assay with brain resident cells, such as neurons, astrocytes, and microglia (Fig. 1a–d and Supplementary Fig. 1a–d). Differentiated SH-SY5Y (dSY5Y) neuronal cells, human and mouse primary astrocytes, and BV2 microglia were stimulated with either a Tolllike receptor 2 (TLR2) agonist or a TLR4 agonist (LPS, 0.1 μg/ml) for 1 h and incubated with α-syn fibrils (200 nM) (Fig. 1a and Supplementary Fig. 1a)
Stimulation of TLR2 for 8, 24, and 2 h resulted in significantly higher levels of α-syn fibril uptake in neurons, astrocytes, and microglial cells (Fig. 1b–d); the levels of α-syn fibril uptake were not altered by TLR4 stimulation (Supplementary Fig. 1b–d)

Summary

Introduction

Introduction Synucleinopathies such asParkinson’s disease (PD) and dementia with Lewy bodies (DLB) are the second most common age-related neurodegenerative disorders and are characterized by abnormal deposits of α-synuclein (α-syn) called Lewy bodies (LBs) and Lewy neurites (LNs)1–3. αSyn pathology spreads through various brain regions during the progression of the disease[4]. A small amount of monomeric and oligomeric α-syn can be released from neurons into the extracellular space and can be taken up by neighboring neurons and glial cells, thereby inducing neurotoxicity[1,6]. We previously demonstrated that the pattern recognition receptor Tolllike receptor 2 (TLR2) can directly interact with β-sheetenriched oligomeric forms of neuron-released α-syn to induce proinflammatory microglial activation[9,10]. Another innate immune receptor, TLR4, is known to play a role in α-syn clearance, and studies have shown that genetic depletion of TLR4 ameliorates neurodegeneration, neuroinflammation, and behavioral deficits in toxin-based mouse models of PD11–14. Additional studies employing recombinant α-syn aggregates of various forms have identified several other α-syn receptors, such as lymphocyte-activation gene 3 (LAG3), cluster of differentiation 36 (CD36), macrophage-1 antigen (MAC-1), nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, β1-integrin, and heparan sulfate[15,16,17,18,19]

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