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Continual loss of cardiomyocytes after myocardial infarction (MI) increases the risk of heart failure., Since cardiomyocyte differentiation is one of the characteristics of bone marrow stromal cells (BMSCs), these cells have been used to repair damaged myocardium in pre-clinical studies and recent clinical trials. However, the treatment effect in humans is modest, in contrast to that in animals, where the effect is marked and significant. Thus, it is necessary to know the mechanism underlying successful cell therapy to resolve the efficacy discrepancy between species. The efficacy of cell therapy is known to be associated with the recruitment of stem cells and chemokines to the injured organ., One critical chemokine is stromal-derived factor 1 (SDF-1), the first chemokine found to link stem cell recruitment and post-MI cardiac repair.
Stromal-derived factor 1 is crucial for directing stem cell migration along a low to high SDF-1 gradient. In the bone marrow, SDF-1 is produced by BMSCs to anchor haematopoietic stem cells. When SDF-1 is degraded by proteases, a gradient between bone marrow and peripheral blood is established. This gradient promotes stem cell migration into the systemic circulation. The subsequent fate of circulating stem cells is determined by another SDF-1 gradient from the peripheral blood to the damaged tissue. For instance, immediately after MI, not only does the concentration of SDF-1 in the myocardium increase, but the number of stem cells in the infarcted myocardium also increases. However, this over-expression of SDF-1 in the myocardium is only maintained for 1 week and the concentration of myocardial SDF-1 is decreased in ischaemic cardiomyopathy. Thus, in chronic MI, an SDF-1 gradient that is unfavourable to stem cell recruitment from blood to remodelled myocardium may occur.
Taken together, an SDF-1 gradient from bone marrow to blood and from blood to the myocardium may play a critical role in stem cell recruitment into the heart. Importantly, BMSCs are cells that can secrete high concentrations of SDF-1. Thus, the beneficial effects of BMSC-based cell therapy could be associated with the change in the SDF-1 gradient. In this study, using rabbits as an experimental model of chronic MI, we determined whether BMSC transplantation could recruit more stem cells to the heart in order to improve left ventricular (LV) remodelling, and we explored the changes in SDF-1 levels in the bone marrow, peripheral blood, and myocardium before and after cell therapy.
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The main findings of our study are that BMSC-based cell therapy increased the SDF-1 gradient from bone marrow to peripheral blood and from peripheral blood to the myocardium, BMSC-based cell therapy increased the number of stem cells in the remodelled myocardium, and BMSC-based cell therapy significantly improved LV remodelling with respect to infarct size, capillary density, and cardiac function. These results suggest that reversal of the SDF-1 gradient towards the myocardium is a novel effect of cell therapy, which acts to recruit more stem cells for cardiac repair.
Interestingly, we observed that SDF-1 levels in both bone marrow and peripheral blood tended to be increased in rabbits with chronic MI. One possible explanation for the increase of marrow SDF-1 is that persistent coronary ligation increases myocardial apoptosis, leading to an increase in DNA damage which in turn causes a significant increase in SDF-1 in the bone marrow. On the other hand, in line with previous reports, we found that myocardial SDF-1 levels decreased in chronic MI. A cellular loss after MI could partly explain the decrease in myocardial SDF-1 in the infarcted heart because the majority of heart cells constitutively expresses SDF-1. Thus, our data illustrate that an unfavourable SDF-1 gradient for stem cell mobilization from bone marrow to the heart exists in chronic MI.
Despite the unfavourable SDF-1 gradient in chronic MI, we found that EGFP-labelled BMSCs were retained and detected in both the remote and infarcted myocardium at 1 day and 4 weeks after LV intracavitary injection. Indeed, SDF-1 is crucial for directing stem cell migration. However, adhesion of circulating BMSCs in the heart appears to be an endothelium dependent process and is enhanced by TNF-alpha and IL-1beta. In chronic MI, the levels of these cytokines are increased in the non-infarcted remote myocardium. Thus, infused BMSCs can end up in the myocardium, especially in the non-infarcted myocardium as shown in Figure 1. These findings support the feasibility of incorporating cell therapy in the treatment of chronic MI.
In our present study using a rabbit model of chronic MI, the levels of SDF-1 in all parts of the hearts of rabbits receiving BMSCs were higher than those in rabbits receiving saline treatment. Similarly, in a rat model of acute MI, a study by Tang et al. also demonstrated that SDF-1 increases in the BMSC-treated rat hearts compared to media-treated hearts 3 weeks after MI. This increase in myocardial SDF-1 is not an unexpected finding; in our study, the transplanted BMSCs actually engrafted into the myocardium, and BMSCs can produce high levels of SDF-1. In addition, as shown in the present study, preservation or regeneration of cardiomyocytes and an increase in capillary density due to cell therapy may also contribute to the increase in myocardial SDF-1 levels. On the other hand, after cell therapy, the SDF-1 level in the bone marrow is decreased, whereas the SDF-1 level in peripheral blood is elevated, creating an increasing SDF-1 gradient from bone marrow towards the peripheral blood. A possible explanation for the decrease of SDF-1 levels in bone marrow may be that BMSC transplantation reduces myocardial apoptosis and thus decreases the response of the bone marrow to DNA damage and the over-expression of SDF-1. Moreover, before haematopoietic stem cells mobilize from the bone marrow they are divided and differentiated into neutrophils. When excessive stem cells mobilize, active neutrophils accumulate leading to an accumulation of neutrophil proteases able to directly cleave SDF-1 in the bone marrow.
The goal of cell therapy in chronic MI is to optimize LV remodelling and regenerate myocardial structures. The ability of cardiac MRI to assess the extent of scar tissue in the myocardium with high spatial resolution is one of the major strengths of the present study. We found that along with the three- to four-fold increase in the number of stem cells, the infarct scar size decreased by 34% at 1 month after cell therapy. This observation supports the notion that transplanted BMSCs or recruited stem cells participate in the myocardial regeneration or cardiomyocyte preservation in infarcted rabbits. Furthermore, our results demonstrate that myocardial SDF-1 levels increase after cell therapy. SDF-1 has been shown to promote survival of BMSCs and cardiomyocytes. Thus, the increase in myocardial SDF-1 might also play a role in improving LV remodelling.
The next question is whether these findings in rabbits are applicable to humans. Based on our study, it is conceivable that cell therapy may be more effective in improving LV function if the cell source is BMSCs and if an adequate number of BMSCs is given. Compared to haematopoietic stem cells, BMSCs are a better source for cell therapy because they are more easily expanded in culture and produce more SDF-1. To date, in most clinical studies, investigators were concerned about the risk of contamination occurring during the in vitro expansion step and used unselected bone marrow cells or bone marrow mononuclear cells as the source of cell therapy. A recent meta-analysis of these studies showed an increase in mean LVEF of 4%. In contrast, animal studies have mainly used BMSCs and have shown an increase in mean LVEF of 8% in MI models in various species with cell doses of 0.06–4 × 106 cells/kg body weight., Bone marrow mononuclear cells contain an extremely low number of BMSCs (1/104–1/105). Thus, even though clinical investigators transplanted a total mononuclear cell number of up to 108, they actually delivered less than 104 BMSCs (or 170 cells /kg). In our study with infarcted rabbits, a delivery of 2 × 106 BMSCs (0.7 × 106 cells/kg) into rabbit was enough to change the SDF-1 gradient and improve cardiac remodelling. Considering the difference in BMSC amount between rabbits and humans, we speculate that a dose of 106 BMSCs/kg may be sufficient for humans. Supporting this notion, Chen et al. administered high doses of BMSCs (108 cells/kg) in patients with recent MI. At 6 months, there was an absolute increase in LVEF of 18% in the BMSC group. Further studies measuring cell dose-responses would allow us to quantitatively compare the outcomes and suggest the optimal dose for human studies.
In summary, this study demonstrates that BMSC-based cell therapy generates an SDF-1 gradient towards the heart, concurrently recruit more stem cells to the heart, and improves LV remodelling. Our results suggest that this novel effect of cell therapy could occur in post-infarction patients if adequate numbers of BMSCs were transplanted.