Cultures of oral epithelial cells can be generated to confluence on AM expanded ex vivo from biopsy-derived oral mucosal tissue. Autologous transplantation was performed with these cultivated oral epithelial cells onto the ocular surfaces of keratectomized rabbit eyes. Autologous transplantation of cultivated oral epithelium is a feasible method for ocular surface reconstruction. The long-term outcome of such transplantation is not yet clear, and its feasibility in clinical use should be evaluated further.

Abstract-Matrix metalloproteinases (MMPs) have been implicated in the process of neovascularization. However, the exact roles of individual MMPs in vessel formation are poorly understood. To study the putative role of MMP-2 in ischemia-induced neovascularization, a hindlimb ischemia model was applied to MMP-2 ϩ/ϩ and MMP-2 Ϫ/Ϫ mice. Serial laser Doppler blood-flow analysis revealed that the recovery of the ischemic/normal blood-flow ratio in MMP-2 Ϫ/Ϫ young and old mice remained impaired throughout the follow-up period. At day 35, microangiography and anti-L-lectin immunohistochemical staining revealed lesser developed collateral vessels and capillary formation in both old and young MMP-2 Ϫ/Ϫ mice compared with the age-matched MMP-2 ϩ/ϩ mice. An aortic-ring culture assay showed a markedly impaired angiogenic response in MMP-2 Ϫ/Ϫ mice, which was partially recovered by supplementation of the culture medium with recombinant MMP-2. Aorta-derived endothelial cells or bone marrow-derived endothelial progenitor cell (EPC)-like c-Kit ϩ cells from MMP-2 Ϫ/Ϫ showed marked impairment of invasive or/and proliferative abilities. At day 7, plasma and ischemic tissues of vascular endothelial growth factor protein were reduced in MMP-2 Ϫ/Ϫ . Flow cytometry showed that the numbers of EPC-like CD31 ϩ c-Kit ϩ cells in peripheral blood markedly decreased in MMP-2-deficient mice. Transplantation of bone marrow-derived mononuclear cells from MMP-2 ϩ/ϩ mice restored neovascularization in MMP-2 Ϫ/Ϫ young mice. These data suggest that MMP-2 deficiency impairs ischemia-induced neovascularization through a reduction of endothelial cell and EPC invasive and/or proliferative activities and EPC mobilization. (Circ Res. 2007;100:904-913.) Key Words: ischemia Ⅲ angiogenesis Ⅲ matrix metalloproteinase Ⅲ endothelium Ⅲ mobilization Ⅲ migration I t is well known that the process of new blood vessel formation is associated with extracellular matrix (ECM) remodeling involving various proteolytic systems. Among such systems, matrix metalloproteinases (MMPs) are a family of zincdependent endopeptidases comprising at least 20 members that are collectively capable of degrading all known ECM components. 1,2 A number of studies have shown that various kinds of MMPs were upregulated in ischemia-induced angiogenesis. 3 Although MMP activity is commonly thought to be involved in the process of angiogenesis, this notion has been challenged by recent studies using genetic or biological target methods. It has been reported that MMP-9 deficiency reduced neovascularization and tumor growth. 4 Study of membrane-type1 (MT1)-MMP knockout mice revealed that the deficiency impaired neovascularization in a mouse corneal micropocket model. 5 Whereas MMP-1 and MMP-10 appear to control the process of vascular regression rather than morphogenesis. 6 On the other hand, certain MMPs, including MMP-12 and MMP-7, are capable of converting plasminogen into angiostatin to inhibit endothelial cell (EC) tubulogenesis in vitro. 7 Interestingly, it has been reported that tiss...

Background Exercise stimulates the vascular response in pathological conditions, including ischemia; however, the molecular mechanisms by which exercise improves the impaired hypoxia-induced factor (HIF)-1α–mediated response to hypoxia associated with aging are poorly understood. Here, we report that swimming training (ST) modulates the vascular response to ischemia in aged (24-month-old) mice. Methods and Results Aged wild-type mice (MMP-2+/+) that maintained ST (swimming 1 h/d) from day 1 after surgery were randomly assigned to 4 groups that were treated with either vehicle, LY294002, or deferoxamine for 14 days. Mice that were maintained in a sedentary condition served as controls. ST increased blood flow, capillary density, and levels of p-Akt, HIF-1α, vascular endothelial growth factor, Fit-1, and matrix metalloproteinase-2 (MMP-2) in MMP-2+/+ mice. ST also increased the numbers of circulating endothelial progenitor cells and their function associated with activation of HIF-1α. All of these effects were diminished by LY294002, an inhibitor of phosphatidylinositol 3-kinase; enhanced by deferoxamine, an HIF-1α stabilizer; and impaired by knockout of MMP-2. Finally, bone marrow transplantation confirmed that ST enhanced endothelial progenitor cell homing to ischemic sites in aged mice. Conclusions ST can improve neovascularization in response to hypoxia via a phosphatidylinositol 3-kinase–dependent mechanism that is mediated by the HIF-1α/vascular endothelial growth factor/MMP-2 pathway in advanced age.

Recently we demonstrated the occurrence of a novel device of signal transducers in Escherichia coli. This class of bacterial sensory kinases, typified by ArcB and BarA, possesses two phospho-donor (His) sites, together with a phospho-accepting (Asp) site. These multi-phosphorylation sites were suggested to make a phosphotransfer circuit. To clarify this complex circuitry, we carried out a series of in vitro assays involving a set of ArcB mutant proteins which have an amino acid substitution at each putative phosphorylation site (His-292, Asp-576 and His-717). By these in vitro phosphorylation and/or phosphotransfer assays, the followings were assessed: (i) ArcB autophosphorylation; (ii) ArcB-mediated phosphorylation of the cognate response regulator, ArcA; (iii) ArcB-mediated phosphorylation of its truncated form (ArcBc) encompassing only the C-terminal phosphorylation site (His-717); (iv) phosphotransfer from ArcBc to ArcA; and (v) phosphotransfer from ArcBc to ArcB. On the basis of these in vitro results, a complex circuitry was revealed for the signal transducer ArcB. This evidence obtained in vitro supports the view that ArcB can serve as a powerful device for not only propagating multi-signals, but also making up signalling networks, in ways more sophisticated than previously thought.

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