The main new findings of the present study suggest that intravenous delivery of human UCB-derived CD133+ progenitor cells can prevent scar thinning, attenuate systolic dilatation, and improve LV function after MI. The transfused cells were able to migrate, colonize, and survive in the infarcted myocardium. However, the therapeutic effect was independent of transdifferentiation and direct myogenic or angiogenic contribution.
The Rationale for Using UCB CD133+ Progenitor Cells
Human UCB cells are rich in stem and progenitor cells with improved proliferative characteristics [7, 12, 13]. These cells can be easily obtained, can be expanded in vitro [16, 22, 23], have the potential for enhanced self-renewal and angiogenic and myogenic differentiation [7, 23, 24], and can be “banked” for future use. The collection of UCB from as many donors as possible would also increase the likelihood that people from many ethnic groups would be able to find a match. It appears that there is reduced risk of rejection by the recipient's immune system with UCB-derived stem cells . Cord blood progenitor cells are routinely used in patients affected by major hematological disorders as an alternative to bone marrow transplantation for stem cell reconstitution [16, 26]. The use of cord blood may make stem cell transplants available more quickly for patients with severe cases of MI who need the cells as soon as possible.
Murohara et al.  have shown that a greater number of angioblast-like EPCs developed from cord blood MNCs than from the same amount of adult peripheral blood EPCs. Transplanted UCB EPCs augmented ischemia-induced neovascularization and regional blood flow in immunodeficient rats in vivo . Pesce et al.  have shown that isolated human cord blood CD34+ cells injected into ischemic skeletal muscles give rise to endothelial and skeletal muscle cells in mice .
Most recently, three studies have reported the use of human UCB cells to treat MI in rats and mice. Henning et al.  injected 1 × 106 human UCB MNCs (subpopulation was not reported) immediately after coronary artery occlusion in Sprague-Dawley rats. Immunosuppression was not administered. Compared with the control, UCB cells reduced infarct size and LV dilatation and improved ejection fraction . The mechanism by which human UCB MNCs reduced infarct size was not defined, and vessel density in the scar was not reported. In another study, Hirata et al.  injected 2 × 105 UCB CD34+ cells immediately after ligation of the coronary artery in Wistar rats. The rats were treated with immunosuppression FK 506. In agreement with our findings, 4 weeks after MI, FS was better in UCB-treated rats. Injected cells survived in the scar tissue and improved vessel density, and approximately 1% were incorporated into vessel walls. The authors did not report on immune rejection of the implanted cells. In a most recent report, Ma et al.  infused 6 × 106 UCB MNCs (only <1% CD34 cells) in the tail vein of nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice 24 hours after MI. Human DNA was detected in marrow, spleen, and liver of both infarcted and noninfarcted mice up to 3 weeks after cell injection. In the heart, however, hDNA was detected in 10 of 19 MI mice but in none of the mice without MI. Infarct size was smaller in cell-treated mice than in untreated hearts. In cell-treated mice, capillary density in the infarct border zone was approximately 20% higher and clusters of UCB-derived cells were detected in the perivascular interstitium. The majority of neovessels appeared to consist of mouse cells. Notably, up to 70% of the cord blood-derived cells in the heart were CD45+. There was no evidence of cardiomyocyte transdifferentiation. Collectively, our findings add strength to the evidence and extend those findings by showing that i.v. delivery of UCB CD133+ cells, 1 week after MI, is feasible and effective.
Mechanism of Therapeutic Effect of UCB CD133+ Cells
Our findings suggest that the benefit is likely a result of factors secreted by the human-derived cells or another type of interaction with the healing infarct rather than a direct mechanical or angiogenic contribution. The idea that progenitor cells may protect the myocardium without directly participating in myocardial repair is supported by several recent works in cardiac [30, 31] and renal disease [32, 33] models.
The present study suggests at least one mechanism that could explain the functional recovery resulting from UCB cell delivery: It might work through prevention of scar thinning by autologous myofibroblast accumulation. By thickening the scar, wall stress is reduced (Laplace law) as is the degree of outward motion of the infarct that occurs during systole (paradoxical systolic bulging) [34, 35]. As a consequence, although the cells did not influence LV diastolic dilatation and mass, there was a lower end-systolic volume in the treated group . This is a significant effect because one of the most important predictors of mortality in patients with MI is the degree of LV systolic dilatation .
CD45 and CD68 staining suggests that the implanted human cells differentiated toward a hematopoietic lineage. None of the nine hearts in the cell group showed the presence of myofibroblasts expressing HLA-DR. Thus, the myofibroblasts at the scar tissue did not originate from a human source.
The intention of the present study was not to confirm or refute the ability of the cells to transdifferentiate or fuse with cardiomyocytes or vascular cells. Although a few human cells were incorporated into vascular endothelium (suggesting direct contribution to neovascularization), we could not exclude the possibility of cell fusion. Recent reports support the notion that hematopoietic progenitors can differentiate into cardiomyocytes and endothelial cells [37, 38]. Furthermore, the therapeutic effect on the infarcted myocardium could be independent of direct contribution to cardiac parenchyma, either myocardial or vascular in nature . Other reports have suggested that the therapeutic effects of cells on LV remodeling and function might be independent of implanted cell survival [30, 31], trans-differentiation [15, 30, 31], scar vessel density [31, 39], or infarct size .
We are aware of several limitations in our study. Based on our previous study , we delivered the cells intravenously 1 week after MI. Homing of the CD133+ progenitor cells into the infarcted heart is allied to the expression of the stem cell chemokine stromal-derived growth factor-1 (SDF-1) in the infarcted tissue and its CXCR4 receptor on human progenitor cells [40, 41]. However, SDF-1 gene expression is highest in the first days after infarction [40, 42]. Thus, it is possible that earlier cell delivery could achieve better migration and therapeutic effect. However, it has been suggested that increased levels of several genes in addition to SDF-1, including those for vascular endothelial growth factor (VEGF), intercellular adhesion molecule-1, vascular cell adhesion molecule-1, stem cell factor (SCF), and hepatocyte growth factor (HGF), might act in concert with SDF-1 to recruit progenitors to the injured heart [43, 44]. We chose to deliver cells 7 days after MI to avoid cell loss due to intense inflammation and washout at the infarct zone. The time of cell delivery, in the present study, might be more relevant to the clinical scenario of cell therapy for MI patients and allow baseline evaluation for myocardial damage and viability as well as donor and recipient HLA matching.
We and other groups described the body distribution of stem cells after systemic i.v. delivery [19, 29, 45, 46]. Heart failure after extensive MI produces congestion and subsequent injury in several organs, including spleen, lungs, liver, kidney, and gut. Tissue damage leads to a dramatic increase in the levels of secreted chemokines, cytokines, and proteolytic enzymes in many organs as part of the regeneration and repair process which have profound impacts on stem cell migration and repopulation . It is possible that hypoxia induced factor-1 (HIF-1) activation and a shift in SDF-1 expression by endothelial cells in other organs during stress influence stem cell homing in different directions . Taken together, the cells distribute and colonize widely to a variety of nonhematopoietic tissues following systemic infusion after MI and may possess the capacity to proliferate within these tissues [46, 47]. The significance of this phenomenon, which also exists in human patients , is uncertain and needs further research. Finally, the duration of the beneficial effect on heart is unknown and might be transient .