Figure 1
Stimulation of EphB2 induces tau dephosphorylation in cells and hippocampus of human tau transgenic mice.
(a) SK-N-SH but not HEK293-tau cells express endogenous EphB2 receptor measured by Western blotting. (b, c) EphB2 was stimulated by treatment of the SK-N-SH cells with ephrinB1/Fc dissolved in DMEM for 45 min and then tau phosphorylation at different sites as labeled was measured by Western blotting. (d, e) The flag-labeled wild type EphB2 (EphB2wt) plasmid was transfected into HEK293-tau cells for 24 h and then the cells were treated with ephrinB1/Fc for 45 min before detection of tau phosphorylation. (f, g) The primary hippocampal neurons (cultured for 7 days in vitro) were treated with ephrinB1/Fc for 45 min and then tau phosphorylation was measured by Western blotting. (h, i) The ephrinB1/Fc was infused into hippocampal CA3 region of the human tau transgenic mice (10 m old) for 45 min and then tau phosphorylation in hippocampus was measured by immunohistochemistry staining (h) and quantitative analysis (i). Scale bar = 20 μm. Dephosphorylation of tau was detected by stimulation of EphB2 receptor. The total tau was probed by R134d or Tau5 that was normalized against tubulin (DM1A) and the phosphorylation level of tau at Thr205, Thr231, Ser396 and tau-1 sites was normalized against total tau. [Note that tau-1 reacts with the unphosphorylated tau at Ser198/199/202, therefore an increased immunoreaction to tau-1 suggests an increased tau dephosphorylation]. The blots were representative of at least three independent experiments with cells from 9 ~ 12 culture wells for each group. The images were representative of at least 3 mice. *P < 0.05, **P < 0.01 versus Fc or DMEM or vector; #P < 0.05 versus EphB2 wt alone (mean±SD).
