Fig. 4. Fasudil regulated ROCK2/moesin/ABCG2 on membrane of TMZ-R cells.

a IP studies were performed with p-ROCK2 and moesin using U251 and U251R. b U251R and rG-1 cells were treated with fasudil (10 μM) for 24 h, and IP was then performed to detect the interaction between p-ROCK2 and moesin. c The interaction of moesin and ABCG2 was detected by IP in U251R and rG-1 cells. d U251R and rG-1 cells were treated with fasudil (10 μM) for 24 h, and IP was then performed to detect the interaction between moesin and ABCG2. e, f U251R and rG-1 cells were treated with XAV939 (10 μM) for 8 h, and then cells were stimulated with fasudil for 2 and 4 h, and western blot was performed for ABCG2 expression. g U251 cell was transfected with ABCG2 overexpression plasmid, and ABCG2 protein was determined by western blot. h Transfected with ABCG2 plasmid, expression of ABCG2 of membrane (M) and total (T) was detected at different time points (0, 4, 8, 12, 16 and 24 h) in U251 cell. i U251 cell was transfected with ABCG2 overexpression plasmid, then fasudil (10 μM) was used to treat cell, and ABCG2 expression of membrane (M) and total (T) was determined at different time points (0, 4, 8, 12, 16, 24 h). j Transfected with ABCG2 plasmid, expression of ABCG2 of membrane (M) and total (T) was detected at different time points (0, 4, 8, 12, 16, 24 h) in U251 cell under treatment of LPA (10 μM). k U251 cell was transfected with ABCG2 overexpression plasmid, then cell was treated with LPA (10 μM) and fasudil (10 μM), ABCG2 expression of membrane (M) and total (T) was determined at different time points (0, 4, 8, 12, 16, 24 h). GAPDH and flotillin-1 were used as loading control