GA prevents BSCB disruption by attenuating MMP-9 expression and activation after SCI
We next asked whether GA treatment inhibits the expression and activity of MMP-2 and MMP-9, which contribute to the disruption of B-BB/BSCB. As shown in Fig. 4A, the level of Mmp-9 mRNA was increased after injury compared with that of the sham control. However, the SCI-induced increase in Mmp-9 mRNA expression was significantly inhibited by GA treatment at 6 h and 1 d after injury as compared with vehicle control, whereas Mmp-2 mRNA expression was not. By gelatin zymography, both the pro-form MMP-2 and MMP-9 were increased at 1 d after injury, but the active form was increased only in MMP-9, as in the previous reports (Lee et al., 2014b; Lee et al., 2015; Lee et al., 2018). Furthermore, both pro-form MMP-9 and active-form MMP-9 were significantly inhibited by GA as compared with vehicle control, indicating that GA treatment inhibits MMP-9 activity after SCI (Fig. 4B and 4C).
Next, we determined the effect of GA on the BSCB permeability by an Evans blue assay on the control or GA-treated spinal cord at 1 d after SCI. As shown in Fig. 4D, the amount of Evans blue dye extravasation was increased after SCI when compared with that of the sham control, which indicates that BSCB disruption was induced after SCI. Furthermore, GA treatment significantly reduced the amount of Evans blue dye extravasation, depending on the dose, after SCI (Fig. 4D and E). We next examined the effect of GA on the tight junction (TJ) protein levels of ZO-1 and occludin at 1 d and 5 d after injury. As shown in Fig. 4F, the decrease of the levels of ZO-1 at 1 d and occludin at 5 d after SCI was significantly attenuated by GA treatment as compared with vehicle control.