Figure 6

Aβ modulated Per2 expression. (A) Temporal expression of Per2 mRNA in Aβ-treated HT22 cell. Per2 mRNA levels in Aβ-treated HT22 cells were decreased and disrupted by circadian oscillation compared to vehicle-treated HT22 cells. Total RNA of HT22 cells was prepared at 4 h intervals. The mRNA level of Per2 was quantified by real-time PCR. Data were represented as mean ± SEM. ^* P < 0.05, ^** P < 0.01, ^*** P < 0.001. (B) HT22 cells were transiently transfected with Per2-luciferacse promoter construct. HT22 cells were treated with vehicle or Aβ (2 μM) for CT12, 24 after cell synchronization. Per2-promoter activity was significantly decreased by Aβ treatment. Data are represented as mean ± SEM. ^** P < 0.01. (C) HT22 cells were incubated with vehicle or Aβ (2 μM) for CT12, 24 after cell synchronization. Western blot analysis revealed that PER2 protein expression was decreased by Aβ treatment at CT24. (D) Densitometric analysis of PER protein from three independent experiments. PER2 was normalized to GAPDH. Data were represented as mean ± SEM. ^* P < 0.05. (E) Photomicrograph of immunostaining with PER2 in HT22 cells shows that PER2 immunofluorescence was significantly decreased by 2 μM of Aβ for 24 h. (F) Densitometric analysis of PER2 immunofluorescence from three independent experiments. Data were represented as mean ± SEM. ^** P < 0.01. (G) HT22 cells were treated Aβ with both L685,458 and siSumo1 for CT12, CT24 after cell synchronization. PER2 expression was significantly increased by L685,458 and siSumo1 treatment. (H) Densitometric analysis of PER protein from three independent experiments. PER2 was normalized to GAPDH. Data are represented as mean ± SEM. ^** P < 0.01, ^## P < 0.01, ^# P < 0.05.