Figure 3.
Ifnb–/– Mice Develop LBs with α-syn, Ubiquitin, and pTau
(A) IF of α-syn in SN of 6-month-old Ifnb–/– mice. Scale bar, 10 μm; 2 μm in close-up lower panel.
(B) IF of 3-month-old Ifnb–/– brains showing α-syn+ neuron in frontal cortex and positive aggregates in STR. Scale bars, 10 μm.
(C) IHC of α-syn in thalamus. Scale bar, 20 μm.
(D) Number of α-syn-positive LBs in thalamus of 6- and 12-month-old mice. ∗∗∗p < 0.001 by Mann-Whitney U test for two groups, †††p < 0.001 by Kruskal-Wallis test for all groups.
(E–G) (E) WB of α-syn and quantified IOD in (F) soluble and (G) insoluble BG fractions of 1.5-month-old mice. Data are mean ± SEM, n = 4.
(H and I) (H) WB of α-syn and (I) quantified IOD in TX-100 soluble and insoluble fractions of 3-month-old brains. Data are mean ± SEM, n = 5.
(J) RT-PCR of α-syn in Ifnb–/– relative to Ifnb+/+ brains. Data are mean ± SEM, n = 3–5.
(K) WB of α-syn (TX-100 soluble and insoluble) from BG of 8-month-old mice and long exposure of whole membranes of soluble fractions.
(L and M) Quantified IOD of α-syn and pSer129-α-syn in (L) insoluble and (M) soluble fractions of monomeric (short exposure) and high-molecular-weight α-syn species (long exposure). Graphs, mean ± SEM, n = 5.
(N) WB and quantified ubiquitin IOD in BG from 1.5-month-old mice. Data are mean ± SEM, n = 3.
(O) IHC of ubiquitin in SN at 3 months and quantified ubiquitin+ aggregates/μm2. Scale bar, 200 μm; 20 μm in inserts. Graph, mean ± SEM, n = 5.
(P and Q) IHC of ubiquitin in locus coeruleus at (P) 3 months and (Q) 12 months. Scale bar, 200 μm.
(R) WB and quantified ubiquitin IOD in brain extracts from 3-month-old mice. Data are mean ± SEM, n = 3.
(S and T) IF and quantification of (S) ubiquitin+α-syn+ and (T) p-tau+α-syn+ aggregates in thalami of 12-month-old mice. Scale bar, 10 μm; data are percentage double positives ± SEM.
(U) IHC of α-syn in mice thalami showing LB-like structures (arrows). Scale bar, 50 μm. Graph, α-syn+ LB-like structures in thalamus ± SEM, n = 5. ∗∗∗p < 0.001 by one-way ANOVA and Turkey’s post hoc correction test.
(V and W) Immuno-EM of thalamic neurons from Ifnb–/– and Ifnar–/– mice showing LB-like structures positive for α-syn by (V) immuno-DAB reactivity and (W) immunogold labeling.
For (I), (L), (M), (O), (R), (S), and (T), ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 by Student’s t test. See also Figure S3.