Improving the engraftment and integration of cell transplantation for cardiac regeneration

This editorial refers to ‘N-cadherin overexpression enhances the reparative potency of human-induced pluripotent stem cell-derived cardiac myocytes in infarcted mouse hearts’, by X. Lou et al., pp. 671–685.

Myocardial infarction (MI), or heart attack, is a common cardiac condition caused by the clogging of the coronary artery that leads to the deprivation of nutrients and oxygen in the heart tissue. In an event of MI, patients may lose up to 1 billion cardiomyocytes (CMs). Unfortunately, adult CMs have a limited capacity for self-renewal, insufficient to compensate for the damage caused by MI. Consequently, cardiac functional deterioration after MI is largely irreversible. Indeed, MI patients often face a dire prognosis, with one study estimating that the 1-year mortality rate is 10-fold higher in the MI patients than the general population.¹

Extensive efforts have been dedicated to regenerating the heart after MI. Cell therapy, in particular, has emerged as one of the important approaches. To date, numerous different cell types have been explored with varying degrees of success in both preclinical and clinical studies, including pluripotent stem cell (PSC)-derived CMs, mesenchymal stem cells (MSCs), bone marrow progenitor cells, and cardiosphere cells.² Transplanted cells (e.g. MSCs) may release soluble factors or exosomes to reduce inflammation³ and promote angiogenesis,⁴ hence minimizing the injury and promoting recovery. In addition, transplanted cells, which now include PSC-derived CMs, can also directly contribute to the formation of new cardiac tissue.⁵

However, despite the promising progress, major obstacles remain to be tackled in this field. For instance, the efficacy of cell transplantation is often crippled by its low engraftment rate. This issue is especially problematic in the case of cardiac regeneration. Specifically, MI creates a harsh microenvironment with elevated oxidative stress along with limited nutrients and oxygen supply, inducing damage and apoptosis in the delivered cells. One animal study estimates that only 5% of embryonic stem cell-derived CMs were grafted 4 weeks after injection.⁵ Similarly, another study found that a mere 2% of MSCs managed to survive after myocardial delivery.⁶ In addition to engraftment, the success of CM transplantation also requires functional integration with the host tissue, as the electrical de-coupling between exogenous CMs and host myocardium raises the risk of arrhythmias.

Lou et al. describe a novel method to improve the engraftment and functional integration of human-induced pluripotent stem cell-derived CMs (hiPSC-CMs) in a mouse MI model.⁷ Specifically, hiPSCs were transfected with an αMHC-CDH2 plasmid. As a result, these genetically engineered cells overexpress CDH2 as they differentiate into CMs (αMHC-positive cells). The rationale behind this strategy is three-fold. First, CDH2, also known as N-Cadherin, plays a vital role in cell–cell interaction and is required for the electro-mechanical synchronization of cardiac tissue.⁸ Second, it was previously shown that CDH2 regulates the activity of PI3K/AKT signalling pathway⁹ and hence its overexpression may promote cell survival. Third, CDH2 has been shown to stimulate angiogenesis,¹⁰ a process critical for tissue regeneration. Taken together, the authors hypothesize that CDH2-overexpressing hiPSC-CMs (CDH2-hiPSC-CMs) may have enhanced therapeutic potential.

To test the hypothesis, the authors performed in vitro characterization of these engineered cells and found significant functional improvements. Specifically, cell–cell interaction in the CDH2-hiPSC-CMs was improved. In particular, these cells had a higher expression of gap junction protein Connexin 43, a molecule critical for cardiac electrical coupling. Indeed, optical mapping showed these cells also had a faster conduction velocity than their wild type (WT) counterparts. In addition, CDH2-hiPSC-CMs exhibited elevated resistance towards hypoxia-induced apoptosis. This result hints that CDH2 cells may better survive the infarcted heart tissue than normal CMs. Mechanistically, the authors attributed enhanced survival to activation of AKT pathway by CDH2 as indicated by AKT phosphorylation.

Subsequent in vivo study also yielded promising results. Upon induction of MI in the mouse, CDH2-hiPSC-CMs or WT-hiPSC-CMs were injected into the infarct regions. In alignment with the in vitro data, CDH2 cells showed a two to three times greater engraftment rate than the WT cells as evidenced by bioluminescence imaging and histology evaluation. Interestingly, CDH2 cells also induced a stronger vasculogenic response around the peri-infarct zone as indicated by the immunostaining of isolectin B4, an endothelial marker. This angiogenic effect was partially attributed to increased secretion of VEGF in the CDH2 cells. More importantly, the increased engraftment rate and angiogenesis translated into more effective cardiac regeneration. Twenty-eight days after MI, mice treated with CDH2 cells or WT cells had a 60% and a 40% reduction in infarct size, respectively, compared with untreated mice. Additionally, echocardiography showed that ejection fraction (EF) was better preserved in the CDH2 group (50%) than the WT group (40%), whereas EF of the untreated group dropped to 20%. Taken together, this study presented solid evidence supporting that CDH2 overexpression is a promising strategy to improve the outcome of cell therapy.

In the past two decades, cell therapy has emerged as a highly attractive strategy for cardiac regeneration. A wide variety of methods have been developed in order to improve the engraftment and integration of delivered cells into the host tissue, such as pretreating the cells with pro-survival cocktails,¹¹ application of pro-survival biomaterials,¹² hypoxia preconditioning,¹³ and various tissue engineering techniques¹⁴ (Figure 1). This study by Lou et al. provides a valuable new addition to this expanding toolkit. Nevertheless, despite these rapid advancements, many roadblocks remain before cell therapy can be safely and widely adopted for cardiac regeneration in humans. For instance, PSC-derived CMs may be functionally and structurally distinct from the host myocardium due to their immaturity, posing an increased risk for electrical de-coupling and arrhythmias. Injection of PSC-derived cells also raises a safety concern for teratoma formation due to the potent proliferation of undifferentiated PSCs.¹⁵ Furthermore, common animal models of MI using mice, rats, and guinea-pigs have fundamentally different characteristics from humans, casting doubt on the validity of these models. Lastly, the exact mechanism underlying the benefits of cell therapy is still controversial.

Figure 1.

Summary of strategies to improve cell engraftment and integration in cardiac regeneration after MI. Methods include pretreatment of cells with pro-survival cocktails, hypoxia preconditioning, engineered hydrogel for encapsulation, genetic modifications of the cells, and co-delivery of growth factors or oxygen using nano- and microparticles.

In conclusion, cell therapy for cardiac regeneration is a fledgling field with tremendous potentials as well as formidable challenges. Undoubtedly, there is still a long way to go before we can widely translate this technology into clinical applications. In future studies, we must leverage multidisciplinary collaborations among biologists, biomedical engineers, material scientists, and clinicians in order to design an effective and safe protocol to regenerate human hearts.

The opinions expressed in this article are not necessarily those of the Editors of Cardiovascular Research or of the European Society of Cardiology.