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.