α-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells

Christian Hansen; Elodie Angot; Ann-Louise Bergström; Jennifer A. Steiner; Laura Pieri; Gesine Paul; Tiago F. Outeiro; Ronald Melki; Pekka Kallunki; Karina Fog; Jia-Yi Li; Patrik Brundin

doi:10.1172/JCI43366

Figure 3

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Intercellular propagation of α-syn in SH-SY5Y cell culture.

(A) Representative picture showing double-labeled SH-SY5Y cells after 14 days of coculture of stable SH-SY5Y cell lines expressing α-syn fused to either GFP or DsRed. Arrows represent transferred aggregates. (B) Confocal microscopy: representative pictures showing double-labeled SH-SY5Y cells after 14 days of coculture of stable SH-SY5Y cell lines expressing α-syn fused to either GFP or DsRed. Scale bars: 2 μM. (C) Upper panel: representative pictures showing double-labeled PC12 cells after 4 days of coculture of human SKMel5 and rat PC12/GFP cells. α-Syn was detected with a monoclonal antibody specific for human α-syn and an Alexa Fluor 568–coupled secondary antibody directed against mouse. Lower panel: comparative Western blot showing the expression level of α-syn in PC12 and SKMel5 cell lysates. Arrows represent double-labeled cells. (D) Representative pictures showing double-labeled N2A cells after 7 days of coculture of human SKMel5 and mouse N2A cells. α-Syn was detected with a monoclonal antibody specific for human α-syn and a Cy3-coupled secondary antibody directed against mouse, and N2A cells were probed with an antibody specific for MAP2 and an Alexa Fluor 488–coupled secondary antibody directed against rabbit. Arrows represent double-labeled cells. Original magnification, ×100 (A); ×40 (C); ×60 (D).