Adenovirus-induced Reactive Astrogliosis Exacerbates the Pathology of Parkinson's Disease

Abstract

Parkinson's disease (PD) is the most prevalent neurodegenerative motor disorder. While PD has been attributed to dopaminergic neuronal death in substantia nigra pars compacta (SNpc), accumulating lines of evidence have suggested that reactive astrogliosis is critically involved in PD pathology. These pathological changes are associated with α-synuclein aggregation, which is more prone to be induced by an A53T mutation. Therefore, the overexpression of A53T-mutated α-synuclein (A53T-α-syn) has been utilized as a popular animal model of PD. However, this animal model only shows marginal-to-moderate extents of reactive astrogliosis and astrocytic α-synuclein accumulation, while these phenomena are prominent in human PD brains. Here we show that Adeno-GFAP-GFP virus injection into SNpc causes severe reactive astrogliosis and exacerbates the A53T-α-syn-mediated PD pathology. In particular, we demonstrate that AAV-CMV-A53T-α-syn injection, when combined with Adeno-GFAP-GFP, causes more significant loss of dopaminergic neuronal tyrosine hydroxylase level and gain of astrocytic GFAP and GABA levels. Moreover, the combination of AAV-CMV-A53T-α-syn and Adeno-GFAP-GFP causes an extensive astrocytic α-syn expression, just as in human PD brains. These results are in marked contrast to previous reports that AAV-CMV-A53T-α-syn alone causes α-syn expression mostly in neurons but rarely in astrocytes. Furthermore, the combination causes a severe PD-like motor dysfunction as assessed by rotarod and cylinder tests within three weeks from the virus injection, whereas Adeno-GFAP-GFP alone or AAV-CMV-A53T-α-syn alone does not. Our findings implicate that inducing reactive astrogliosis exacerbates PD-like pathologies and propose the virus combination as an advanced strategy for developing a new animal model of PD.

Keywords: Adenovirus infections; Alpha-synuclein; Mouse model; Parkinson’s disease; Reactive astrogliosis.

Fig. 1

Adenovirus injection exacerbates A53T-mediated parkinsonian motor deficits. (a) Schematic diagram of virus injection (left) and experimental schedule (right). (b) Quantification of latency-to-fall assessed by rotarod test. (c) Quantification of asymmetry score of forelimb usage assessed by cylinder test. For all figures, mean±SEM; ns, non-significance; *p<0.05, **p<0.01, ***p<0.001 assessed by Kruskal-Wallis test with Dunn's multiple comparison test (b) or One-way ANOVA with Tukey’s multiple comparison test (c).

Fig. 2

Adenovirus injection exacerbates A53T-mediated TH loss. (a) Representative images of TH-stained SNpc tissues (left) and striatal tissues (right). (b) Quantification of TH-positive (TH+) neurons in the SNpc (at the AP coordinates of -2.7~3.0, -3.0~-3.3, -3.3~-3.6, -3.6~-4.0 mm). (c) Quantification of TH optical density in the striatum. For all figures, mean±SEM; ns, non-significance; *p<0.05, **p<0.01, ***p<0.001 assessed by Kruskal-Wallis test with Dunn’s multiple comparison test (b; AP -3.0~-3.3) or One-way ANOVA with Tukey’s multiple comparison test.

Fig. 3

Adenovirus injection exacerbates reactive astrogliosis. (a) Representative confocal images of SNpc tissues stained with TH, GFAP, and GABA. (b) Quantification of the volume of GFAP-positive (GFAP⁺) astrocytes in the SNpc. (c) Quantification of the integrated density of GABA in the GFAP-positive astrocytes. For all figures, mean±SEM; ns, non-significance, ***p<0.001 assessed by Kruskal-Wallis test with Dunn's multiple comparison test.

Fig. 4

Adenovirus injection exacerbates the astrocytic expression of α-syn. (a) Representative confocal images of SNpc tissues stained with TH, GFAP, and α-syn. Yellow arrowheads indicate the neuronal α-syn expression, while white arrowheads indicate the astrocytic α-syn expression. (b) Quantification of α-syn immunoreactivity in neurons. (c) Quantification of α-syn immunoreactivity in the GFAP-positive (GFAP⁺) astrocytes. For all figures, mean±SEM; ns, non-significance; *p<0.05, **p<0.01, ***p<0.001 assessed by Kruskal-Wallis test with Dunn's multiple comparison test.