The San Diego, California-based biotechnology company, CardioCell, has sponsored a preclinical study in laboratory mice that explored giving preconditioned mesenchymal stem cells intravenously to animals that had suffered large heart attacks. This study “Mesenchymal stem cells grown under chronic hypoxia traffic to regions of myocardial infarction, suppress splenic natural killer cells, and attenuate adverse remodeling in mice with large acute MI” at the European Society of Cardiology (ESC) Congress by one of the authors, Dr. Michael Lipinski, Interventional Cardiologist at MedStar Washington Hospital Center. Lipinski collaborated with Drs. Dror Luger, Research Scientist at Washington Hospital Center and Stephen Epstein, Director, Translational and Vascular Biology Research at MedStar Heart and Vascular Institute, Chair of CardioCell’s Scientific Advisory Board and Member of CardioCell’s Heart Failure Advisory Board.
The vast majority of stem cell-based studies to treat heart attacks have examined stem cell injections directly into the heart muscle or using techniques associated with stent placement to administer stem cells through the blood vessels that surround the heart. Stem cell injection directly into the heart muscle requires special equipment and personnel who have special training. This procedure also carries the risk of rupture of the wall of the heart, even though skilled practitioners can reduce such risks. Intracoronary administration of stem cells does not require specialized training or equipment, since any cardiologist can perform such a procedure. However, several studies have shown that the vast majority of the stem cells administered in this fashion end up in the lung. Intravenous administration would potentially be the safest and easiest way to administer such stem cells. The problem with intravenous (IV) administration of stem cells is that no one has been able to show that IV administration of stem cells works for heart attack patients.
The problem is that IV stem cells do not know where to go. However, some IV-administered stem cells do find their way to the damaged heart. How can we make the majority of such stem cells go to where they are so badly needed?
Several experiments have examined ways to do this. In particular, mesenchymal stem cells (MSCs) move towards increasing concentrations of a small protein called “stromal cell-derived factor-1” (SDF-1). The receptor for SDF-1, CXCR4 (a member of the Src family of protein kinases, for those who are interested) binds SDF-1, and when it does so, it kicks the cell in the rump, wakes it up, and drives toward higher and higher concentrations of SDF-1. Several laboratories have forced the expression of higher levels of CXCR4 in stem cells in order to improve their ability to home to damaged tissues, because damaged tissues express SDF-1. Such a strategy works (one example, see Cheng, M., et al., J Mol Cell Cardiol. 2015;81:49-53), and even increases tissue healing, but increasing CXCR4 levels in stem cells without resorting to genetic engineering techniques is preferable, since such a procedure would not only be safer and less expensive, it would require fewer regulatory hurdles to pass FDA muster (see Park JS, et al., Methods. 2015;84:3-16).
CardioCell’s new preclinical study has examined the ability of ischemia-tolerant mesenchymal stem cells (itMSCs), when administered intravenously, to heal the heart after a heart attack. They examined the ability of IV administration of itMSCs to prevent the deterioration of the left ventricle and the onset of “remodeling” that occurs after a heart attack (i.e. “remodeling” refers to the enlargement of the heart that leads to heart failure). Secondly, this study examined the ability of itMSCs to reduce the inflammation that develops after a heart attack and responsible for the continued death of heart muscle and heart blood vessel cells.
It has been well established that itMSCs secrete factors that have marked abilities to staunch inflammation. Therefore this study was specifically designed to determine if intravenously administered itMSCs can improve cardiac function following a heart attack and if such improvements were mediated by systemic anti-inflammatory activities. According to Stephen Epstein, “The study impressively demonstrates the validity of these concepts. IV itMSC administration indeed improves cardiac function, and the itMSCs achieve this – at least, in part, – by their anti-inflammatory effects and abilities to decrease NK cells. These findings can profoundly impact future strategies for treating patients with AMI.”
For this study, CD1 male mice were given heart attacks by surgically occluding the left anterior descending artery (LAD). After surgery, these mice were given infusions of itMSCs into their tail veins. These itMSCs were continuously grown at 5% O2. Stem cell infusions were given 24 hours following the experimentally-induced heart attacks. The infused stem cells were labeled with a radioisotope (indium-111 oxine), which allowed them to be easily visualized in the bodies of the mice. Each animal was infused with one million cells. So-called “ex vivo phosphor” imaging of the heart was performed 24 hours following itMSC injection.
In a separate study, mice were subjected to baseline echocardiography followed by surgery in which they were given heart attacks. Then 24 hours later, the mice were randomized to two groups, one of which received an infusion of two million itMSCs and the other of which received infusions of physiological saline solution. There were 16 mice in both groups.
Echocardiography was repeated at 3, 7 and 21 days after the induction of the heart attack. Blood, spleen and hearts were then harvested and the heart were stained with TTC staining of the hearts — TTC staining identifies dead versus live tissue. Live tissue stains a deep red color and dead tissue appears white.
Radiolabeled itMSCs preferentially homed to regions of myocardial injury. However, it must be admitted that the total number of the injected cells that engrafted in the myocardium was small. There was minimal homing in control mice that had been given stem cell infusions but were not given heart attacks. The size of the infarction was no different in the two groups as ascertained by TTC staining (28±3% for itMSC group vs. 25±3% for control group). As expected, control saline-treated mice with large infarcts that covered more than 25% of the left ventricle showed an increase in adverse compared to mice with smaller infarcts. However, mice treated with itMSCs did not demonstrate an increase in adverse remodeling, regardless of the size of their infarcts. This seems to demonstrate that itMSCs prevented the adverse LV remodeling occurring in mice with large infarcts.
Additionally, the heart wall thickness were greater in the itMSC group both during contraction and relaxation compared with the control group. Importantly, itMSC injection caused a significant decrease in splenic Natural Killer cells compared with control injection (2.6±0.13 vs. 3.4±0.36, p<0.04). This is important, because Natural Killer (NK) cells play a major role in post-heart attack inflammation and cause to good deal of the damage to the heart after a heart attack. Other experiments in cell culture showed that itMSCs significantly suppress NK cell proliferation as a consequence of the cocktail of molecules they secrete.
This study was predicated upon and entirely different strategy than the paradigms that have driven previous stem cell experiments. The vast majority of previous stem cell experiments assumed that implanted stem cells improve cardiac outcomes by regenerating heart tissue or by stimulating endogenous stem cell populations to engraft into the myocardium and regenerate the damaged heart. Thus, the greater the number of stem cells that engraft into the heart, the better.
In this study, a different paradigm was being tested. In this case, IV delivery results in very low numbers of cells engrafting into the damaged myocardium. However, the stem cells are not expected to contribute to myocardial regeneration; instead they are expected to stem the excessive immune or inflammatory responses that cause the progressive deterioration of the heart that dogs heart attack patients. In this study, the authors wished to examine if IV administered itMSCs grown under chronic hypoxic conditions could improve myocardial function and adverse remodeling, and if a functional benefit occurs, does the anti-inflammatory effects of itMSCs play an important mechanistic role in this improvement. In this study, both hypotheses seem to be valid.
CardioCell’s itMSCs secrete higher levels of growth factors and other important proteins associated with neoangiogenesis and healing and also seem to home to damaged tissues better than MSCs grown under normal conditions. This preclinical study might provide fodder for further experiments and discussions in the months and years to come.