Effects of ent -Sauchinone on STAT3 DNA binding activity and protein expressions of STAT3 were detected by western blotting using specific antibodies in astrocytes and microglial BV-2 cells. Astrocytes (A) and microglial BV-2 cells (B) were treated with 1 mg/ml of LPS alone or with LPS plus different concentrations (1, 5, and 10 μM) of ent-Sauchinone at 37°C for 1 hour. DNA binding activity of STAT1 and STAT3 was investigated using EMSA as described in Materials and Methods. Nuclear extracts were subjected to DNA binding reaction with 32P end-labeled oligonucleotide specific to STAT3. Specific DNA binding of the STAT3 complex is indicated by an arrow. Similar results were treated with 1 μg/ml of LPS alone, or with LPS plus different concentrations (1, 5, and 10 μM) of ent-Sauchinone at 37°C for 1 hour. Equal amounts of total proteins (40 μg/lane) were subjected to 10% SDS-PAGE, and activation of STAT3 (phosphorylation) was detected by western blotting using specific antibodies in astrocytes (C) and in microglial BV-2 cells (D). Values below or above of each figures mean quantified relative expression of the proteins or DNA binding activity.