Fig. 5
From: Pathological α-synuclein recruits LRRK2 expressing pro-inflammatory monocytes to the brain
Increased α-synuclein fibril-induced infiltration of pro-inflammatory monocytes in Lrrk2 knock-in R1441C and G2019S-BAC mice. A Representative immunoblots and quantification of Lrrk2 expression and Rab10 phosphorylation (pT73-Rab10 to total Rab10) in brain lysates from WT (nTg) mice and homozygous Lrrk2-R1441C knock-in (KI) mice, and B live CD45hi/CD11b+/Ly6Chi midbrain cells isolated from a mononuclear-enriched Percoll gradient 3 days after injection (bilaterally) with 10 μg of human α-synuclein fibrils (gating strategy is presented in Supplemental Fig. 8). C, D Mice expressing G2019S-Lrrk2 with the mutation knocked into a mouse BAC spanning the Lrrk2 gene were likewise analyzed together with littermate WT (nTg) controls. E Quantification of CD45hi/CD11b+/Ly6Chi (enriched in pro-inflammatory classical monocytes) cells isolated from the brains of WT (nTg), Lrrk2-R1441C KI and G2019S BAC mice 72 h after the injection of 10 μg (bilaterally) of monomeric human α-synuclein. F In contrast to monomeric protein injections, the injection of fibrils (10 μg bilateral, 72 h wait) results in the robust recruitment of CD45hi/CD11b+/Ly6Chi cells in the mouse brain (PBS perfused and then analyzed by flow cytometry, see Methods) that is further increased in R1441C-KI mice and G G2019S-BAC mice. Microglia cell numbers after fibril treatment between strains are not different between groups (see Supplemental Fig. 8). H Percentage of activated microglia (CD45lo/CD11b+/MHCII+) in WT (nTg) and R1441C-KI mice injected with saline, human α-synuclein monomers, or fibrils (10 μg bilateral) is shown in the graph. In each panel, each dot represents the analysis of an individual mouse with 13 mice (3 males and 10 females) analyzed in E, 22 mice (9 males and 13 females) analyzed for F, 14 mice (5 males and 9 females) analyzed for G, and 37 mice (13 males and 24 females) analyzed for H. All mice were aged 2–3 months at the time of the analysis. Data are plotted as the mean (red bars) ± SEM. Significance is determined by unpaired t-tests *p < 0.05, **p < 0.01