Abstract 226: Mechanisms Underlying the Impairment of Ischemia-induced Neovascularization in Aging Mice: the Pivotal Role for Matirx-Metalloproteinase-2
Background: It has been reported that neovascularization was impaired in aging humans and animals, and matrix metalloproteinases (MMPs) have been implicated in this process. However, the exact roles of individual MMPs in vessel formation in the aging are poorly understood.
Methods and Results: To study the aging-dependent role of MMP-2 in ischemia-induced neovascularization, we applied a hindlimb ischemia model to young (6 weeks of age) and old (72 weeks of age) MMP-2+/+ and MMP-2−/− mice. Serial Laser Doppler blood-flow analysis revealed that the recovery of the ischemic-normal blood-flow ratio was lower by 59% in old MMP-2+/+ than in young MMP-2+/+ (n = 10, p < 0.01) and by 35% in old MMP-2−/− (n= 10, p < 0.01) than in old MMP-2+/+ at 14 days after ischemia. At day 28, microangiography and immunohistostaining revealed lesser-developed collateral vessels and capillary formation in old MMP-2−/− compared to the relative age-matched controls. An aortic-ring culture assay showed a markedly impaired angiogenic response in the old MMP-2−/− aorta, which was recovered by supplementation of the culture medium with recombinant MMP-2 (2.8-fold, n = 6; p < 0.001), although to a lesser extent than in the young MMP-2+/+ aorta (5.1-fold, n = 6; p < 0.0001). MMP-2 deficiency dramatically impaired the invasive ability of aorta-derived endothelial cells (ECs) from old MMP-2−/− (67%, n = 6; p < 0.001). Moreover, MMP-2 deficiency reduced not only the levels of ischemic muscles in old MMP-2−/− (43%, n = 6, p < 0.05) but also the level of MMP-2 on the EC surface stimulated with VEGF (32%, n = 5, p < 0.01). At 10 days after ischemia, the numbers of EPC-like CD31+c-Kit+ mononuclear cells (MNCs) in peripheral blood were decreased in MMP-2−/−, though the change was not significant. Transplantation of bone-marrow (BM) MNCs from MMP-2+/+ young mice partially restored neovascularization in old MMP-2−/− (35%, n = 8), but restored neovascularization did not result from transplantation of old MMP-2+/+ BM-derived MNCs.
Conclusions: These data suggest that aging impairs ischemia-induced neovascularization, and that the mechanisms of this impairment in old MMP-2−/− may be mediated in part by impairment of EC invasive activity via aging-dependent reduction of MMP-2 expression on the EC surface.