Research Paper Volume 16, Issue 10 pp 9168—9187

Figure 2. Stress hormone activates DNA damage-Chk1-CIP2A pathway, results in tau hyperphosphorylation, Aβ overproduction and synaptic impairments in primary neurons. (A–E) Primary neurons were treated with 100 nM corticotropin-releasing factor (CRF) for 24 hours. (A) Representative immunoblots of γH2A.X, Chk1-pS317, Chk1-pS345, Chk1, CIP2A, Tau-pT231, Tau-pS396, Tau-pS404, Tau-5 (total tau), APP-pT668, APP and β-actin. (B) Quantification of the relative protein levels; non-phosphorylated proteins such as γH2A.X and CIP2A were normalized to the β-actin levels; phosphorylated Chk1-pS317, Chk1-pS345, Tau-pT231, Tau-pS396, Tau-pS404 and APP-pT668 were normalized to corresponding total Chk1, tau (Tau-5) and APP respectively. n = 3 or n = 6 per group. (C) The relative concentration of Aβ₄₀ and Aβ₄₂ in supernatant and cell lysate of primary neurons detected by ELISA. n = 3 or n = 4 per group. (D) Representative immunoblots of Synapsin I, PSD95, GluA1 and β-actin. (E) Quantification of the relative protein levels, which were normalized to the β-actin levels. n = 3 per group. Data are presented as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001.