Abstract 1479: Fibrin(ogen) Stimulates Bone Marrow-Derived Endothelial Progenitor Cell Survival and Proliferation: Implications for Pro-Angiogenic Cell Therapy
Bone marrow-mononuclear cells (BMC) can be used as a source of endothelial progenitor cells (EPC) to enhance post-ischemic neovascularization by cell therapy. There is growing interest in methods to expand EPC prior to delivery. Expanding EPC without compromising their differentiation potential or survival would allow to increase therapeutic preparations, reduce the required size of donor tissue, homogenize the cell phenotype, and possibly facilitate pretreatments to augment the therapeutic efficacy of the cell preparations. We hypothesized that extracellular matrices (ECM) modulate BMC pro-angiogenic efficacy. We expanded BMC on various substrates: Plastic, plastic-immobilized purified fibrinogen, fibronectin, gelatin-vitronectin, or gels of fibrin or Matrigel. BMC cultured on fibrinogen for 2–3 weeks produced confluent endothelial-like cobblestone cell monolayers, unlike any other ECM tested. Fibrinogen and fibrin increased BMC differentiation into EPC by 57,9% compared to gelatin-vitronectin (p<0.05), as shown by UEA-1 lectin binding and Dil-acetylated LDL incorporation. Cell cycle analysis by FACS showed that fibrinogen stimulated proliferation by 84.5% over gelatin-vitronectin after 3 d (p<0.05), and conferred high resistance to cell death in response to apoptotic challenges such as ceramide, staurosporine or etoposide. In a mouse model of hindlimb ischemia after femoral artery ligation, injection of BMC cultured onto fibrinogen for 3 d enhanced neovascularization by 38.2% over BMC cultured on gelatin-vitronectin (p<0.05), as measured by Laser-Doppler foot perfusion imaging, microangiographic scoring and evaluation of capillary density. In summary, we showed that fibrin(ogen) stimulates BMC proliferation, survival and differentiation into EPC in vitro, and enhanced their pro-angiogenic effects in cell therapy. Fibrinogen thus appears an ideal ECM to expand EPC on prior to cell therapy. Culture on fibrinogen may be useful prior to transplantation of EPC with compromised angiogenic function in conditions such as diabetes, obesity or atherosclerosis. In addition, our data suggest that fibrin(ogen) deposition may stimulate EPC-mediated tissue neo-vascularization in conditions associated with thrombosis.