Background. We demonstrated previously that intracoronary (IC) injection, in a dose-escalating fashion, of autologous bone marrow (BM)-derived mononuclear cells (MNCs) in normal pigs (n = 3) was safe. Although IC injection with the highest cell dose (15 x 10^6 cells) resulted in transient significantly reduced [Thrombolysis In Myocardial Infarction (TIMI) grade 1] flow in the injected artery, this did not result in myocardial infarction (MI), as determined by measurement of plasma biomarkers of myocardial necrosis and histopathologic examination. We further studied IC injection of autologous BM-derived MNCs in a porcine chronic myocardial ischemia model. Here we report the biodistribution of CM dioctadecyl tetramethylindocarbocyanine (CM-DiI)-labeled MNCs in solid organs and BM following IC injection.
Methods. Chronic myocardial ischemia was induced in domestic pigs (n = 5) by surgical placement via lateral thoracotomy of an ameroid cuff around the distal left circumflex artery. Coronary angiography three weeks later confirmed absent or significantly reduced (TIMI grades 0 or 1, respectively) coronary blood flow. BM was aspirated (40 – 60 mL, mean 45.4 mL) from the right femur. MNCs were isolated by Ficoll density gradient centrifugation. Cells were labeled with CM-DiI at 37 °C for 20 minutes. A total of 15 x 106 MNCs in 5 mL of phosphate-buffered saline with autologous plasma was injected by IC delivery. Seven days later, BM was aspirated from the left femur, after which animals were euthanized. The heart and major organs were harvested for histological studies. Alternate sections were either stained with hematoxylin and eosin or left unstained for immunofluoroscence microscopy for CM-DiI visualization.
Findings. Neither gross inspection nor histopathology revealed evidence of MI. Cell injection resulted in TIMI grade 1 coronary flow that resolved after 10 minutes and was not associated with vasospasm. There was no significant elevation in plasma biomarkers of myocardial necrosis at one or seven days following cell therapy. A limited number of labeled MNCs were observed in perivascular structures in myocardium at the injection site. Labeled cells were not seen elsewhere in either the myocardium or identified in lung, liver, or kidney. DiI-labeled MNC localization was observed in spleen harvested from one of the animals. BM aspirates, which did not allow for quantitation of cell numbers, indicated labeled MNCs.
Conclusions. IC injection of DiI-labeled BM-derived MNCs resulted in cell localization in the perivascular areas of myocardium supplied by the injected vessel. Cell localization was not observed in most major solid organs, with spleen localization identified in only one animal. Labeled cells were identified in BM aspirates from the three animals injected with CM-DiI-labeled MNCs, suggesting a role for BM engraftment and repopulation as a possible mechanism for MNC localization in myocardium. It remains to be determined whether IC MNC injection resulted in initial homing to BM followed by migration to myocardium, or whether IC cell delivery resulted in initial myocardial deposition followed by BM localization. Ongoing studies include additional biodistribution studies of DiI-labeled BM- and umbilical cord blood-derived stem cell populations in a rat model of acute MI.