Effect of baicalin on liver fatty acid binding protein in oxidative stress model in vitro

Objective To investigate the effect of baicalin on liver fatty acid binding protein in oxidative stress model in vitro.Methods (1) Cellular oxidative stress in vitro was induced by incubating cells with 400μmol/L hydrogen peroxide (H2O2) for 20 minutes at 37℃ in the dark.After Chang liver cell line was treated with different dose of baicalin for 24,48 and 72 hours.MTT assay was employed to detect cell viability,and then the hydrogen peroxide (TC50) of the different dose of baicalin was calculated.(2) Based on MTT assay,cells were treated with three different doses of baicalin (25,50,100 μmol/L) for 24 and 48 hours before being exposed to 400 μmol/L H2O2 for 20 minutes at 37℃.Then,reactive oxygen species (ROS) assay and activity assays of superoxide dismutase (SOD) and reduced glutathione hormone (GSH) were evaluated.(3) Realtime PCR and Western blotting were applied to explore the influence of baicalin on the expression level of L-FABP.(4) One-way ANOVA was used for results statistical analysis.Reesult (1) MTT assay showed baicalin treatment at 25,50,100 μmol/L for 24 and 48 hours was feasible (83.60％ ± 3.47％,72.36％ ±2.18％,70.16％ ± 2.04％ for 24 hour; 84.93％ ± 3.11％o,76.16％ ± 2.45％,72.72％ ± 2.31％ for 48 hours,P ＞ 0.05,F=386.24,475.92 respectively).Meanwhile,we found by the linear regression model that the median toxic concentration of baicalin for 48 hours was 170.6 μmol/L,and the median toxic concentration of baicalin for 24 hours was 153.2 μmol/L.(2) ROS assay showed dichlorofluorescin in all baicalin-treated cells after stress was significantly reduced (37.0 ± 3.30,22.90 ± 3.84,29.60 ± 2.52 for 24 hours respectively,P ＜ 0.05,F =70.06; 35.77 ± 2.35,21.80 ± 3.10,23.87 ± 1.98 for 48 hours respectively,P ＜ 0.05,F =110.92) as compared with the H2O2-treated cells.Moreover,50 μmol/L baicalin treatment for 48 hours was the optimal condition against ROS generation (21.80 ± 3.10,P ＜ 0.01,F =110.92).Furthermore,the activities of intracellular SOD and GSH was increased significantly (51.53 ± 1.91 μg/mg for SOD,P＜ 0.05,F =93.81; 49.85 ± 1.45 U/mg for GSH,P ＜ 0.05,F =92.51).(3) Although realtime PCR analysis indicated 50 μmol/L baicalin treatment for 48 hours could have no changes of the level of L-FABP expression under the oxidative stress condition,western blotting analysis indicated 50 μmol/L baicalin treatment for 48 hours could increase up to about 80％ for the level of L-FABP expression.Conclusion Baicalin was suggested to be able to enhance both L-FABP expression and activity of intracellular SOD and GSH,and therefore protected hepatocytes from oxidative stress. Key words: Reactive oxygen species;  Oxidative stress;  Baicalin;  Liver fatty acid binding protein