Fig. 6

Inhibition of MAPK pathways abrogates LPA-induced ROS and NO production and decreases microglial neurotoxicity. a A ROS assay kit was used to determine changes in the ROS levels generated by primary murine microglia. Serum-starved cells were treated with DMSO and DMSO plus LPA (1 μM) in the absence or presence of SP600125 (10 μM), SB203580 (10 μM), or PD98059 (10 μM) for 30 min and 24 h, followed by 30 min incubation with carboxy-H₂DCFDA, and the fluorescence intensity was evaluated. Results from three experiments are presented as mean values ± SD. RLU, relative luminsescence units. b Serum-starved microglia cells were incubated with vehicle DMSO and DMSO plus LPA (1 μM) in the absence or presence of SP600125 (10 μM), SB203580 (10 μM), or PD98059 (10 μM) for the indicated time periods, and the production of NO was determined by measuring the total nitrate concentration in the supernatants. Data (4 separate experiments) are presented as mean values ± SD. c CATH.a neurons were incubated for 24 h with conditioned media collected from LPA-treated primary microglia in the absence or presence of SP600125 (10 μM), SB203580 (10 μM) or PD98059 (10 μM) for the indicated time periods. The LDH levels were detected and cytotoxicity was calculated according to the manufacturer’s recommendations. (*p < 0.05, **p < 0.01, ***p < 0.001 compared to vehicle control; ^#p < 0.05, ^##p < 0.01^###_,p < 0.001 each inhibitor compared to LPA treated cells; two-way ANOVA with Bonferroni correction)