Subtractive Cloning Identifies Tissue Inhibitor of Matrix Metalloproteinase-1 (TIMP-1) Increased Gene Expression Following Focal Stroke

Background and Purpose—Differential gene expression has been reported following the onset of focal stroke. To identify de novo expression of ischemia-induced genes, we applied subtractive cDNA library strategy to identify the genes that are selectively upregulated by focal stroke. Methods—Spontaneously hypertensive rats were subjected to permanent occlusion of the middle cerebral artery (MCAO). mRNAs prepared from ischemic and nonischemic cortex 2 and 12 hours after MCAO were subtracted, and a subtractive cDNA library was constructed. A cDNA that encodes for tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) was identified in the subtractive cDNA library. The temporal expression of cortical TIMP-1 mRNA was further characterized in ischemic cortex subjected to permanent or temporary (160-minute) MCAO. Results—A panel of genes isolated from the subtractive cDNA library was subjected to Southern analysis to confirm ischemia-induced gene expression. TIMP-1 demonstrated robust induction after ischemic inj...