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The translation of cell-based therapies for ischemic tissue repair remains limited by several factors, including poor cell survival and limited target site retention. Advances in nanotechnology enable the development of specifically designed delivery matrices to address these limitations and thereby improve the efficacy of cell-based therapies. Given the relevance of integrin signaling for cellular homeostasis, we developed an injectable, bioactive peptide-based nanofiber matrix that presents an integrin-binding epitope derived from fibronectin, and evaluated its feasibility as a supportive artificial matrix for bone marrow-derived pro-angiogenic cells (BMPACs) used as a therapy in ischemic tissue repair. Incubation of BMPACs with these peptide nanofibers in vitro significantly attenuated apoptosis while enhancing proliferation and adhesion. Pro-angiogenic function was enhanced, as cells readily formed tubes. These effects were, in part, mediated via p38, and p44/p42 MAP kinases, which are downstream pathways of focal adhesion kinase. In a murine model of hind limb ischemia, an intramuscular injection of BMPACs within this bioactive peptide nanofiber matrix resulted in greater retention of cells, enhanced capillary density, increased limb perfusion, reduced necrosis/amputation, and preserved function of the ischemic limb compared to treatment with cells alone. This self-assembling, bioactive peptide nanofiber matrix presenting an integrin-binding domain of fibronectin improves regenerative efficacy of cell-based strategies in ischemic tissue by enhancing cell survival, retention, and reparative functions., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

There is great demand for the development of novel therapies for ischemic cardiovascular disease, a leading cause of morbidity and mortality worldwide. We report here on the development of a completely synthetic cell-free therapy based on peptide amphiphile nanostructures designed to mimic the activity of VEGF, one of the most potent angiogenic signaling proteins. Following self-assembly of peptide amphiphiles, nanoscale filaments form that display on their surfaces a VEGF-mimetic peptide at high density. The VEGF-mimetic filaments were found to induce phosphorylation of VEGF receptors and promote proangiogenic behavior in endothelial cells, indicated by an enhancement in proliferation, survival, and migration in vitro. In a chicken embryo assay, these nanostructures elicited an angiogenic response in the host vasculature. When evaluated in a mouse hind-limb ischemia model, the nanofibers increased tissue perfusion, functional recovery, limb salvage, and treadmill endurance compared to controls, which included the VEGF-mimetic peptide alone. Immunohistological evidence also demonstrated an increase in the density of microcirculation in the ischemic hind limb, suggesting the mechanism of efficacy of this promising potential therapy is linked to the enhanced microcirculatory angiogenesis that results from treatment with these polyvalent VEGF-mimetic nanofibers.

Cell-based therapy has emerged as a promising therapeutic tool for treatment of ischemic cardiovascular disease. Both unselected bone marrow-derived mononuclear cells (BMNCs), which include stem/progenitor cells and several other cell types, and endothelial progenitor cells (EPCs), a subpopulation of BMNCs, display regenerative potential in ischemic tissue. Abundant evidence supports the involvement of EPCs in capillary growth, and EPCs also appear to participate in the formation of collateral vessels. Collectively, these effects have led to improved perfusion and functional recovery in animal models of myocardial and peripheral ischemia, and in early clinical trials, the therapeutic administration of EPCs to patients with myocardial infarction or chronic angina has been associated with positive trends in perfusion. EPCs also contribute to endothelial repair and may, consequently, impede the development or progression of arteriosclerosis. This review provides a brief summary of the preclinical and clinical evidence for the role of EPCs in blood-vessel formation and repair during ischemic cardiovascular disease., (Copyright 2010 Elsevier Inc. All rights reserved.)

Despite significant advances in medical, interventional, and surgical therapy for coronary and peripheral arterial disease, the burden of these illnesses remains high. To address this unmet need, the science of therapeutic angiogenesis has been evolving for almost two decades. Early preclinical studies and phase I clinical trials achieved promising results with growth factors administered as recombinant proteins or as single-agent gene therapies, and data accumulated through 10 years of clinical trials indicate that gene therapy has an acceptable safety profile. However, more rigorous phase II and phase III clinical trials have failed to unequivocally demonstrate that angiogenic agents are beneficial under the conditions and in the patients studied to date. Investigators have worked to understand the biology of the vascular system and to incorporate their findings into new treatments for patients with ischemic disease. Recent gene- and cell-therapy trials have demonstrated the bioactivity of several new agents and treatment strategies. Collectively, these observations have renewed interest in the mechanisms of angiogenesis and deepened our understanding of the complexity of vascular regeneration. Gene therapy that incorporates multiple growth factors, approaches that combine cell and gene therapy, and the administration of "master switch" agents that activate numerous downstream pathways are among the credible and plausible steps forward. In this review, we examine the clinical development of angiogenic gene therapy, summarize several of the lessons learned during the conduct of these trials, and suggest how this prior experience may guide the conduct of future preclinical investigations and clinical trials.

Aims: Neovascularization is an important element of long-term functional recovery during chronic ischaemia. We postulated that haeme oxygenase (HO) is required for progenitor cell recruitment, neovascularization, and blood flow recovery after critical hindlimb ischaemia (HLI)., Methods and Results: The femoral artery was ligated in FVB/N mice proximal to its superficial and deep branches. Blood flow in the ischaemic hindlimb was determined by laser Doppler perfusion imaging. Capillary density was measured by isolectin staining, and mobilization of Sca-1(+)/Kdr(+) progenitor cells by FACS analysis. Progenitor cell recruitment to the ischaemic hindlimb was assessed after Tie2-lacZ transgenic bone marrow transplantation. Blood flow recovery after femoral artery ligation was significantly blunted in mice treated with the HO inhibitor tin protoporphyrin-IX (25 mg/kg i.p., every other day). HO-inhibited mice developed more pronounced limb necrosis, associated with impaired hindlimb motor function. Capillary density in the ischaemic hindlimb and mobilization of Sca-1(+)/Kdr(+) progenitor cells were significantly reduced after HO inhibition. After transplantation of Tie2-lacZ transgenic bone marrow cells into lethally irradiated wild-type mice, fewer LacZ(+) cells were detected in the ischaemic hindlimb muscle of HO-inhibited mice. Mechanistically, HO inhibition prevented the establishment of a stromal cell-derived factor-1 gradient for progenitor cell mobilization between the ischaemic hindlimb and bone marrow., Conclusion: HOs are required for progenitor cell recruitment, neovascularization, and functional recovery after HLI.

The article presents a study on the function of Gli3, a Hedgehog (Hh) signaling transcription factor, in angiogenesis and ischemic tissue repair. It cites the findings of the assessment of capillary density and left ventricular ejection fractions in Gli3-haploinsufficient (Gli3 +/-) mice and their wild-type littermates. It concluded that Gli3 contributes to vessel growth under ischemic and nonischemic situation and modulates angiogenesis and endothelial cell activity in adult mammals.

Endothelial progenitor cells (EPCs) are a heterogeneous subpopulation of bone marrow mononuclear cells that have an enhanced potential for differentiation within the endothelial cell lineage. In response to ischemic injury, EPCs are mobilized from the bone marrow to the peripheral circulation and home to the sites of new vessel growth, where they become incorporated into the growing vasculature. Thus, EPCs can be therapeutically useful for treating ischemic injury or for delivering anti-cancer agents to tumors. [Copyright &y& Elsevier]

Abstract: The morphogen Sonic hedgehog (Shh) promotes neovascularization in adults by inducing pro-angiogenic cytokine expression in fibroblasts; however, the direct effects of Shh on endothelial cell (EC) function during angiogenesis are unknown. Our findings indicate that Shh promotes capillary morphogenesis (tube length on Matrigel™ increased to 271±50% of the length in untreated cells, p =0.00003), induces EC migration (modified Boyden chamber assay, 191±35% of migration in untreated cells, p =0.00009), and increases EC expression of matrix metalloproteinase 9 (MMP-9) and osteopontin (OPN) mRNA (real-time RT-PCR), which are essential for Shh-induced angiogenesis both in vitro and in vivo. Shh activity in ECs is mediated by Rho, rather than through the “classic” Shh signaling pathway, which involves the Gli transcription factors. The Rho dependence of Shh-induced EC angiogenic activity was documented both in vitro, with dominant-negative RhoA and Rho kinase (ROCK) constructs, and in vivo, with the ROCK inhibitor Y27632 in the mouse corneal angiogenesis model. Finally, experiments performed in MMP-9- and OPN-knockout mice confirmed the roles of the ROCK downstream targets MMP-9 and OPN in Shh-induced angiogenesis. Collectively, our results identify a “nonclassical” pathway by which Shh directly modulates EC phenotype and angiogenic activity. [ABSTRACT FROM AUTHOR]

Background—CXC-chemokine receptor 4 (CXCR4) regulates the retention of stem/progenitor cells in the bone marrow (BM), and the CXCR4 antagonist AMD3100 improves recovery from coronary ligation injury by mobilizing stem/ progenitor cells from the BM to the peripheral blood. Thus, we investigated whether AMD3100 also improves recovery from ischemia/reperfusion injury, which more closely mimics myocardial infarction in patients, because blood flow is only temporarily obstructed. Methods and Results—Mice were treated with single subcutaneous injections of AMD3100 (5 mg/kg) or saline after ischemia/reperfusion injury. Three days later, histological measurements of the ratio of infarct area to area at risk were smaller in AMD3100-treated mice than in mice administered saline, and echocardiographic measurements of left ventricular function were greater in the AMD3100-treated mice at week 4. CXCR4+ cells were mobilized for just 1 day in both groups, but the mobilization of scal +/flk1+ cells endured for 7 days in AMD3100-treated mice compared with just 1 day in the saline-treated mice. AMD3100 upregulated BM levels of endothelial nitric oxide synthase (eNOS) and 2 targets of eNOS signaling, matrix metalloproteinase-9 and soluble Kit ligand. Furthermore, the loss of BM eNOS expression abolished the benefit of AMD3100 on scal+/flk1+ cell mobilization without altering the mobilization of CXCR4+ cells, and the cardioprotective effects of AMD3100 were retained in eNOS-knockout mice that had been transplanted with BM from wild-type mice but not in wild-type mice with eNOS-knockout BM. Conclusions—AMD3100 prolongs BM progenitor mobilization and improves recovery from ischemia/reperfusion injury, and these benefits appear to occur through a previously unidentified link between AMD3100 and BM eNOS expression. [ABSTRACT FROM AUTHOR]