Abstract 380: Cell Isolation Procedures Matter: A Comparison of Different Isolation Protocols of Bone Marrow Mononuclear Cells Used for Cell Therapy in Patients with Acute Myocardial Infarction
Recent clinical trials using intracoronary progenitor cell administration in patients with myocardial infarction demonstrated controversial results regarding the neovascularisation and cardiac function. Whereas the REPAIR-AMI trial showed a significant improvement of ejection fraction, no change was detected in the ASTAMI trial. In order to investigate whether different cell isolation protocols might contribute to the different results, we compared the functional activity of bone marrow-derived mononuclear cells (BMC) isolated by the protocols of the above mentioned trials. In the REPAIR-AMI trial, BMC were isolated by Ficoll gradient centrifugation followed by storage at room temperature in x-vivo medium and 20 % autologous serum. In the ASTAMI protocol, Lymphoprep gradient centrifugation was performed and BMC were stored at 4°C in 0.9% NaCl and 20% heparin-plasma. The recovery of total cell number, colony forming unit capacitiy (CFU), and number of mesenchymal stem cells was significantly reduced to 77±4%, 83±16% and 65±15%, respectively, when using the ASTAMI protocol. The capacity of the isolated BMC to migrate in response to SDF-1 was profoundly impaired when using the ASTAMI isolation procedure (healthy BMC: 42±8%, CAD BMC: 78±3% reduction). Total CXCR4 expressing BMC were reduced using the ASTAMI protocol (75±6% vs. 55±8%). In addition, the mean expression of the CXCR4-receptor was reduced to 57±13% after overnight-incubation in the ASTAMI-protocol. Infusion of identical numbers of BMC into the hind limb ischemia model demonstrated a significantly lower recovery of blood flow by BMC isolated by the ASTAMI protocol (57±5% reduction). The use of NaCl and plasma for cell storage and the reduction of the CXCR4-receptor seem to be the major factors of functional impairment of the BMC. Cell isolation protocols have a major impact on the functional activity of cells isolated for clinical cell therapy. The assessment of cell number and CFU capacity may not entirely reflect the functional capacity of cells in vivo. Additional functional testing by using migration assays and particularly in vivo monitoring of cell function in animal models appears to be mandatory to assure proper cell function before embarking on clinical cell therapy trials.