In a porcine model of acute MI with late reperfusion and autologous cardiac cell therapy, we quantified biodistribution using a radiolabeling technique. Most of the BM-MNCs delivered by the intracoronary route were distributed within the heart and lungs. The presence of MI did not modify BM-MNC cardiac homing and engraftment after intracoronary injection. Finally, systemic intravenous BM-MNC injection promoted lung homing rather than cardiac engraftment.
The first prerequisite for cell therapy success is the engraftment and thus, homing of transplanted cells to the target area. Intracoronary delivery offers the advantages of a non-surgical method that can be performed percutaneously during angioplasty for acute MI. In the present study we showed that intracoronary BM-MNC injection can allow cell engraftment into myocardium tissue, whereas peripheral systemic intravenous administration was not efficient at 24 hours. This suggests that the cardiac niche effect favoured by MI was not sufficient to attract intravenously injected BM-MNCs within 24 hours, thus limiting the efficacy of systemic intravenous injection. Chemoattractant factors secreted by the infarcted heart might be too diluted in the body of large animals like pigs to attract BM-MNCs. If this is the case, similar results may be expected in humans. Another hypothesis is that the ability to trap intravenously injected BM-MNCs within the heart is limited, since only 4% to 5% of cardiac output is dedicated to supplying the coronary arteries ). Finally, cardiac engraftment following systemic intravenous BM-MNC delivery might be limited by lung entrapment prior to coronary artery access. Numerous studies have been performed in rodent  and large animal models [13, 14], using intravenous injection of mesenchymal stem cells, showing mesenchymal stem cell engraftment after 24 hours. Several hypotheses can be discussed to explain these discrepancies with our results. First, our protocol closely matched clinical trial protocols, in which a seven-day delay between acute MI and cell injection seems to be the best time for efficacy following intracoronary injection [3, 15]. However, this time may be too long after intravenous cell injection, in regards to local inflammatory response and niche effect. In most studies mesenchymal stem cells were injected 1 to 72 hours after myocardial infarction [12–14]. Second, BM-MNCs were labeled with 99mTechnetium, a radioelement that has a short half life and may not have been detected at 24 hours. However, we did not observe any fluorescent cell within the heart 24 hours after intravenous injection, corroborating our radioactivity data. Finally, mesenchymal stem cells may have a better capability for cardiac homing than BM-MNCs, that contain a very small stem cell number. Interestingly, a recent study comparing intra-aortic, intravenous, and intramyocardial delivery of mesenchymal stem cells in rats observed 5% cell survival at 48 hours after intravenous delivery, mostly in the lungs .
The radioactive labeling using Isolink™ kit did not discriminate cell type, and was not species nor cell specific. In a pilot study we evaluated 99mTc-hexa-methyl-propylen-amine-oxime (HMPAO) labeling, often used in humans. However, 99mTc-HMPAO labeling of BM-MNCs was totally unsuccessful (not shown). We then switched to the radioactive linker Isolink™, which allowed us to inject a large quantity of 99mTc-labeled cells, significantly higher than in other studies [16, 17]. Although, similar to previous studies [11, 18, 19], the viability of Isolink™/99mTc-labeled BM-MNCs was not altered, the adverse effects of labeling on the migratory and functional abilities of BM-MNCs cannot be entirely excluded. The majority of cardiac BM-MNC homing studies were performed in rodents, and only a few studies established the fraction of transplanted cells retained within the myocardium using direct radioactive labeling of the cells. In our study, radioactivity quantification in each organ showed 34.8 ± 9.9% total radioactivity in the heart with an accumulation within the injection site, one hour after intracoronary BM-MNC injection, and 6.0 ± 1.7% at 24 hours. Importantly, histology results were in accordance with scintigraphic imaging data confirming the presence of numerous BM-MNCs at one hour and 24 hours after intracoronary injection. As in our study, Hou et al , using a pig model of reperfused MI with intracoronary cell delivery and radioactive cell quantification, observed a largely right-sided distribution of BM-MNC. However, only 2.6 ± 0.3% of BM-MNCs were detected in the heart, with some BM-MNCs being distributed to the right ventricle. Importantly, the experimental model in this study was xenogeneic, with human BM-MNCs being injected into pigs. Human cells may lack the correct membrane receptors to home into the pig myocardium or may undergo acute lysis due to a xenogeneic reaction. In a recent study, using a similar model of reperfused MI in pigs with autologous BM-MNC transplantation  6.5% of autologous transplanted BM-MNCs were detected in the heart four days after injection, a result close to ours at 24 hours. Finally, cardiac homing cell rate at one hour might be overestimated by including a possible leakage of radioactive label from the cells, although this appears unlikely as no radioactive cell leakage was observed in vitro in the next four hours following radioactive labeling.
Although phase I-II clinical trials have been completed using coronary delivery of BM cells, only one study used coronary delivery of 99mTc-labeled BM-MNCs (in a single patient), and observed intense cardiac cell engraftment . The study of Hofmann et al  showed that in five patients with MI, only 1.3 to 2.6% of 18F-FDG-labeled BM-MNCs were detected in the infarcted myocardium one hour after intracoronary BM-MNC injection. However, the number of injected BM-MNCs was 30-fold higher than in our study, so the absolute number of retained BM-MNCs within the heart was 39 to 78 × 106 BM-MNCs, a number close to ours (34.8 ± 9.9 × 106 retained BM-MNCs). Intracoronary injection of a very high cell number may saturate the binding sites for cardiac cell homing, raising questions about the ideal number of cells to be injected.
After intracoronary injection, we observed a similar biodistribution of BM-MNCs in animals with or without MI, despite the large size of the infarcted area (45% of the left ventricle). Several studies have shown that acute myocardial infarction is followed by an acute local inflammatory reaction involving upregulation of chemokines receptors and adhesion molecules, thereby facilitating adhesion and infiltration of cells involved in tissue repair, including stem cells [21–23]. The dynamic capability of BM-MNCs to migrate and the niche effect are central in regenerative medicine [21–23]. In our model, to carry out intracoronary injection, blood flow was stopped three times for at least two minutes to prevent backflow and prolong contact time between BM-MNCs and myocardium. When BM-MNCs were injected without prior MI induction, each maneuver for intracoronary cell injection resulted in ST-segment elevation, as already described for repeated angioplasty balloon inflations [24, 25]. Although we did not evaluate the intensity of the ischemia induced by balloon inflation, this suggests that the injection technique created local downstream ischemic and preconditioning effects , thus rendering the local microenvironment more receptive to cell homing . A recent study in a pig model of myocardial infarction and intracoronary injection of autologous BM cells, balloon occlusion was found ineffective to increase cell homing . In another study in a similar model, single-bolus delivery was as effective as three balloon-occlusion deliveries . Importantly, both studies were performed only in pigs with myocardial infarction and not in healthy pigs, suggesting that the cardiac niche effect favoured by MI was sufficient to attract injected BM-MNCs without any further effect of balloon occlusion. The fact that the presence and the size of MI did not influence cell engraftment could be useful for cardiac cell therapy in patients with chronic heart failure of non-ischemic origin or with ischemic myocardium without myocardial infarction. Two clinical trials have been published with patients who have had a myocardial infarction at least three months before BM-MNC coronary injection [1, 27, 28]. A clinical benefit was observed in both studies, suggesting that BM-MNCs homed to the myocardium despite the absence of acute myocardial infarction. The hypothesis of cardiac homing in the absence of acute myocardial infarction was recently confirmed in two Phase I cardiac cell therapy clinical trials on patients with chronic ischemic cardiomyopathy and receiving an intracoronary injection of radiolabeled CD133+ or CD34+ cells [16, 29].