Tissue Inhibitor of Matrix Metalloproteinase-3 (TIMP3) Provides Beneficial Effects Post-MI by Promoting Angiogenesis

Introduction: Myocardial infarction (MI) results in loss of cardiomyocytes, adverse extracellular matrix (ECM) remodelling, leading to left ventricular (LV) dilation and dysfunction. Tissue inhibitor of metalloproteinase (TIMPs) are MMP inhibitors, main regulators of ECM integrity. TIMPs can also regulate other aspects of myocardial remodeling such as hypertrophy, fibrosis and inflammation. TIMP3 levels are reduced in the peri-infarct zone within 24 hours post-MI in mice. Hypothesis: Replenishment of TIMP3 post-MI limit infarct expansion, and attenuate LV dilation and dysfunction. Methods: MI was induced in adult male wildtype (C57BL/6) mice by ligation of the left anterior descending artery. Adenoviral constructs expressing human TIMP3 (Ad- hTIMP3) or no-TIMP (Ad-Null, control) were injected in the peri-infarct zone (5.4x10 7 pfu, 5 injections/heart). Cardiac function was assessed by echocardiography. Cardiomyocyte density (WGA/DAPI staining), vascular density (Fluo-lectin injection, CD31 IHC), ECM composition (PSR staining) were assessed at 3 and 7 days post-MI. In vitro, angiogenic potency of TIMP3 (rTIMP3) was assessed using the 3D fibrin gel-based angiogenesis assay using primary human vascular (HUVECs) and coronary artery endothelial cells (HCAECs), and co-IP between TIMP3 and VEGFR2. Results: Ad-TIMP3 injections significantly improved LV function and reduced LV dilation as compared to Ad-Null group post-MI. Infarct size was markedly reduced with TIMP3 injections and more viable myocytes were preserved in the infarct zone at 1wk post-MI. Ad-TIMP3-MI group showed a higher density of endothelial cells and increased coronary density in the infarct and peri-infarct regions compared to the Ad-null group. This suggested that Ad-TIMP3 promotes angiogenesis in the infarcted myocardium. In vitro studies confirmed that rTIMP3 promoted angiogenesis/sprouting in human endothelial cells up to100ng/ml. However at higher concentrations (>1ug/ml), rTIMP3 exerted anti-angiogenic effects by binding to VEGFR2. This function of rTIMP3 appears to be through an MMP-inhibitory mechanism. Conclusion: The novel pro-angiogenic function of TIMP3 post-MI could provide additional beneficial effects in post-MI treatment.