| Matrix metalloproteinases ( MMPs) are one of the most important proteolytic enzymes that degrade the extracellular matrix ( ECM ) . After brain ischemia and reperfusion, there was an increase of MMPs, in particular the activation increase of MMP - 9 and MMP - 2 are associated with vascular permeability, blood - brain barrier ( BBB ) permeability , BBB disruption, inflammatory cell influx and brain edema. The sythetic inhibitor of MMPs is used to block the above changes in different levels. We investigated induction of MMP - 2 and MMP - 9 expression after transient focal cerebral ischemia at protein levels in order to clarify the relation among MMP - 2 , MMP - 9 and tissue injury in ischemic brain.MethodsI use a transient middle cerebral artery ( MAC) occlusion model. The right carotid region was exposed through a cervical midline skin incision and division of the right omohyoid muscle. The external carotid artery ( EGA) was coagulated and divided after trimming the proximal EGA branches. The internal carotid artery (ICA) was then exposed and pterygopalatine artery was ligated with asilk suture . Afterclamping the common and internal carotid arteries, a small incision was made in the EGA and a monofilament suture with a rounded tip was introduced and advanced approximately 18. 5 ±0. 5mm into the ICA through the carotid bifurcation for proximal MACO. Temporal and cellular changes of immunohistochemical MMP - 2 and MMP - 9 expression were compared with different periods If reperfusion from 6hours to 7days after transient MAC occlusion. Coronal sections were cut with 6um thickness and mounted on standard glass slides. Depar-affinvation was achieved by treating the specimens in xylene and subsequent ethano followed by a rinse in PBS. After they were blocked with 10% normal horse serum for 90 minutes. The slides were washed in PBS and incubated with a rabbit polyclonal antibody against MMP -2 and MMP -9 at 1:50 dilution for 16 hours at 25℃ subsequently they were incubated with avidin biotinnorsemdish peroxidase complex for 30 minutes and then developed with the use of DAB as a color substrate. The reaction was stopped by washing the slides in distilited water. To ascertain specific binding of the antibody for the protein, a set of brain sections was stained in similar way with 0. 01 MPBS instead If the first antibody. The values are expressed as x ± s. Significant differences were analyzed using analysis of variance ( ANOVA) and t test.Results(1) At 6 hours, 12 hours of reperfusion after transient MCAO immunocreative MMP -2 became detected in the astrocytes. At 1 day ,2 day after reperfusion the expression of MMP -2 in astrocytes become more intense. At 5day after reperfusion the expression of MMP- 2 had a peak. At 7 day after reperfusion the expression of MMP -2 in macrophages and astrocytes decreased.(2) At 6 hours of reperfusion after transient MCAO immunocr -eative MMP - 9 became detected in the vascular endothelial cells, mi-croglias neutrophils. At 12 hours, 24 hours after reperfusion the expression of MMP - 9 in above cells become intense. At 2day after reperfusion the expression of MMP -9 had a peak. At 5 ~7 day after reperfusion the expression of MMP - 9 in neurons decreased. All a-bove features were associated with their actions in MCAO/R.DiscussionThis study indicated in different time point after MCAO /R the expression of MMP - 2 in astrocytes and macrophages, the expression of MMP - 9 can be detected in endothelial cells, neutrophils, micro-glias and neurons. Immunohistochemical method showed that in normal CNS, MMP -2 and MMP -9 was founded in endothelial cells of brain vascular, astrocytes and microglia, this can explain that in Bonder region there is little expression of MMP - 2 and MMP - 9. Anthony had founded that MMP - 9 can be located in endothelial cell and MMP - 2 can be located in macrophages. This may result from differences in sensitivity of the Immunohistochemical method.This study indicated the expression of MMP - 2 and MMP - 9 was associated w...
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