FIGURE 1.

Schematic representation of the MMP-12 shRNA plasmid construct and mechanism of target gene silencing, experimental design, and timing of the main experimental procedures. (A) The construct consists of the annealed invert repeat MMP-12 shRNA sequences ligated between BamHI and HindIII sites of the pSilencer 4.1-CMV neo expression vector. Following the entry of plasmids into the nucleus of a cell, the CMV promoter drives the formation of the MMP-12 pre-shRNA, a short hairpin molecule specific to MMP-12. The Dicer processes MMP-12 pre-shRNA molecules and the resulting MMP-12 siRNA molecules interact with the target MMP-12 mRNA. This interaction leads to the degradation of MMP-12 mRNA and suppression of MMP-12 gene expression. (B) Rats from both sexes were subjected to transient focal cerebral ischemia for 2 h, followed by reperfusion and various treatments. Rats from the appropriate cohorts were euthanized on various post-reperfusion days, and their brains were processed for real-time PCR analysis. Additional animals from the appropriate cohorts were assessed in various neurological and functional tests at baseline and at regular intervals (post-reperfusion days 1, 3, 5, 7, 14, and 21) after a 2-h focal cerebral ischemia.