Use of Suppression Subtractive Hybridization Strategy for Discovery of Increased Tissue Inhibitor of Matrix Metalloproteinase-1 Gene Expression in Brain Ischemic Tolerance

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Abstract
Brief occlusion of the middle cerebral artery (i.e., ischemic preconditioning; PC) induces significant brain protection to subsequent severe ischemic events. In an effort to discover genes responsible for ischemic tolerance, we have applied a new technique, suppression subtractive hybridization (SSH), to identify genes that are upregulated by PC. Using this SSH approach, a cDNA that encodes tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) was identified. Time course studies using Northern analysis revealed that TIMP-1 mRNA was significantly elevated at 24 hours (3.3-fold over controls, P < 0.05, n = 5) and 2 days (4.3-fold increase, P < 0.01) after PC, corresponding to the onset of significant ischemic tolerance. Our data not only demonstrate the utility of this new polymerase chain reaction—based SSH strategy for discovery of genes differentially expressed in PC, but also suggest a potential role of TIMP-1 in PC-induced ischemic tolerance.