Abstract
Human studies and preclinical models of Parkinson’s disease implicate the involvement of both the innate and adaptive immune systems in disease progression. Further, pro-inflammatory markers are highly enriched near neurons containing pathological forms of alpha synuclein (α-syn), and α-syn overexpression recapitulates neuroinflammatory changes in models of Parkinson’s disease. These data suggest that α-syn may initiate a pathological inflammatory response, however the mechanism by which α-syn initiates neuroinflammation is poorly understood. Silencing endogenous α-syn results in a similar pattern of nigral degeneration observed following α-syn overexpression. Here we aimed to test the hypothesis that loss of α-syn function within nigrostriatal neurons results in neuronal dysfunction, which subsequently stimulates neuroinflammation. Adeno-associated virus (AAV) expressing an short hairpin RNA (shRNA) targeting endogenous α-syn was unilaterally injected into the substantia nigra pars compacta (SNc) of adult rats, after which nigrostriatal pathology and indices of neuroinflammation were examined at 7, 10, 14 and 21 days post-surgery. Removing endogenous α-syn from nigrostriatal neurons resulted in a rapid up-regulation of the major histocompatibility complex class 1 (MHC-1) within transduced nigral neurons. Nigral MHC-1 expression occurred prior to any overt cell death and coincided with the recruitment of reactive microglia and T-cells to affected neurons. Following the induction of neuroinflammation, α-syn knockdown resulted in a 50% loss of nigrostriatal neurons in the SNc and a corresponding loss of nigrostriatal terminals and dopamine (DA) concentrations within the striatum. Expression of a control shRNA did not elicit any pathological changes. Silencing α-syn within glutamatergic neurons of the cerebellum did not elicit inflammation or cell death, suggesting that toxicity initiated by α-syn silencing is specific to DA neurons. These data provide evidence that loss of α-syn function within nigrostriatal neurons initiates a neuronal-mediated neuroinflammatory cascade, involving both the innate and adaptive immune systems, which ultimately results in the death of affected neurons.
Highlights
Our α-syn short hairpin RNA (shRNA) targets a position of the α-syn transcript that does not share sequence homology with either βor γ-synuclein (Supplementary Figure 1A), we aimed to ensure that the α-syn shRNA did not affect expression of β-syn, as this could mask the true effects of α-syn silencing (as has been observed with several α-syn germline knockout mice (Abeliovich et al, 2000; Chandra et al, 2004; Greten-Harrison et al, 2010))
We examined the consequences of α-syn shRNA administration in α-syn germline KO and WT mice. α-syn KO or WT mice received unilateral substantia nigra pars compacta (SNc) injections of associated virus (AAV) expressing the α-syn shRNA or a scrambled control shRNA and tyrosine hydroxylase (TH) positive (TH+) nigral neuron numbers were quantified 1-month later
Expression of the α-syn shRNA significantly decreased TH+ nigral neurons in WT mice (∼45% reduction; Supplementary Figures 1D,E; analysis of variance (ANOVA): F(3,24) = 4.465, p < 0.05), but had no effect on TH+ neurons in α-syn KO mice (Supplementary Figures 1D,F). These data provide further evidence that loss of endogenous α-syn is toxic to nigrostriatal neurons, and combined with previously published controls (Gorbatyuk et al, 2010), exclude the possibility that nigrostriatal cell loss following α-syn shRNA administration is due to non-specific RNA interference (RNAi)-induced toxicity or off-target activity
Summary
A growing body of research suggests that inflammation, mediated by both the innate and adaptive immune systems, is a crucial event in the pathogenesis of Parkinson’s disease (McGeer et al, 1988b; Croisier et al, 2005; Orr et al, 2005; Ouchi et al, 2005; Gerhard et al, 2006; Cebrián et al, 2014b). Within the Parkinson’s disease brain catecholamine neurons express major histocompatibility complex class 1 (MHC-1; McGeer et al, 1988b; Croisier et al, 2005; Orr et al, 2005; Ouchi et al, 2005; Gerhard et al, 2006; Cebrián et al, 2014b), and Lewy bodies and dopamine (DA) neurons label with immunoglobulin G (Orr et al, 2005; Ouchi et al, 2005; Gerhard et al, 2006). Serum or immunoglobulin G isolated from Parkinson’s disease patients can mediate selective destruction of DA neurons in models of Parkinson’s disease (McGeer et al, 1988b; Defazio et al, 1994; Chen et al, 1998; Croisier et al, 2005)
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