Aggregated Alpha‐Synuclein Activates Pro‐Inflammatory NFKB Signaling Pathways Through TLR‐Dependent and Independent Mechanisms in Peripheral Monocytic Cells

Abstract

Toll‐like receptors (TLRs) are pattern recognition molecules that induce inflammatory signaling cascades upon recognition of pathogen or damage associated molecules. In addition to peripheral immune cells including monocytes, TLRs are broadly expressed on cells of the central nervous system (CNS) including microglia, astrocytes, and neurons and their activation induces production of pro‐inflammatory mediators inducing neuroinflammation and neurodegeneration in the chronic state. Prolonged neuroinflammation in the substantia nigra (SN) leading to accumulation of alpha‐synuclein fibril (α‐syn) containing Lewy‐bodies (LBs) and ultimately death of dopaminergic neurons is a pathological hallmark of Parkinson’s disease (PD) and is thought to be a major mechanism driving disease progression. TLRs are an integral component of the feed‐forward neuroinflammatory loop where damage from neurodegeneration generates TLR ligands that further amplify the inflammatory circuit. Importantly, aggregate fibrillar α‐syn can interact with TLRs on microglia in the brain, of which TLR2 and 4 are best characterized; however, the extent to which fibrillar α‐syn interacts with peripheral immune cells including monocytes is poorly understood. Peripheral inflammation may play a causal role in inducing and perpetuating neuroinflammation and accumulation of fibrillar α‐syn has been reported at several peripheral sites including the gut and liver in PD. To model the peripheral immune response to fibrillar α‐syn we exposed THP‐1 monocytic cells in vitro to determine inflammatory signaling pathways altered by α‐syn. Compared to monomeric α‐syn, preformed‐fibrillar (PFF) α‐syn displays overt inflammatory gene upregulation and pathway activation as assessed by RNA‐Seq. Both MyD88 and TRIF mediated TLR signaling pathways were broadly overrepresented in the PFF condition as well as RNAK/RANKL signaling supported by large increases in NFKB signaling and pro‐inflammatory cytokine production including TNF and IL‐1B. These results indicate that α‐syn may act broadly across all TLRs and other pro‐inflammatory receptors such as RANK.