PRECLINICAL STUDY: EDITORIAL COMMENT
Human Stem Cells for Heart Failure TreatmentReady for Prime Time?*
Gianluigi Condorelli, MD, PhD , ,2,* and
Daniele Catalucci, PhD,
Division of Cardiology, Department of Medicine, University of California at San Diego, San Diego, California
Istituto di Ricovero e Cura a Carattere Scientifico "Multimedica," Milan, Italy.
* Reprint requests and correspondence: Dr. Gianluigi Condorelli, Division of Cardiology, Department of Medicine, University of California at San Diego, 9500 Gilman Drive, La Jolla, California 92093. (Email: gcondorelli{at}ucsd.edu).
The possibility of replacing the failing myocardium with newly generated tissue is challenging many researchers in the field of cardiovascular medicine. During the last decade, many types of "stem" cells have been used in an attempt to produce differentiated cardiac myocytes (CMs). Among the various types of putative cardiac progenitor cells, mesenchymal, hematopoietic, and local cardiac cells have been tested for their potential as a source of CM progenitors. However, data obtained using these cells are not univocal. For instance, the highly efficient differentiation rate of hematopoietic (1) and mesenchymal (2) stem cells toward CMs observed in initial studies was not subsequently confirmed (3). Rigorous methodologies, including genetically modified mouse models that permit discrimination between cell fusion and true differentiation, suggested that the few differentiation events after myocardial injection of hematopoietic stem cells (HSCs) could be ascribed to the fusion between pre-existing myocytes and transplanted cells (3). The small yield of HSCs differentiating into CMs may be at the basis of the modest improvement of cardiac function by autologous bone marrow cell injection after myocardial infarction (MI) (4). Moreover, production of growth factors with antiapoptotic effect by injected cells on host myocardium was suggested to be responsible for the slight functional improvement (5,6).
Previous studies have described various types of local mouse and human cardiac stem cells (7), which are still in the process of being characterized. More recently, other sources of differentiated CMs have been described, including spermatogonial cells derived from adult testis (8). Interestingly, the use of spermatogonial cells provides a relevant alternative to embryonic stem cells (ESCs) and may overcome current legal and ethical issues that limit the potential therapeutic use of embryonic cells in many countries.
Mouse embryonic stem cells (mESCs) were originally described 25 years ago (9). Since then, a multitude of well-established cell culture techniques and genetic manipulation with viruses or homologous recombination has been developed. Mouse embryonic stem cells derive from the first few cellular divisions of the zygote (10) and can spontaneously differentiate into beating CMs (11) as well as other cell types. Embryonic stem cells mature in vitro within so-called embryoid bodies (EBs), teratoma-like structures, which contain cells originating from all 3 embryonic layers, namely mesoderm, ectoderm, and endoderm (11). Generation of "bulk" cultures of endothelial stem-CMs has been obtained as a result of a significant increase in the yield of mESC-derived CMs from pluripotent mESCs (12). This improvement has been achieved through genetic selections obtained by transfecting mESCs with a reporter gene containing a drug-resistance marker (an enzyme that metabolizes a toxic drug) under the expression control of a cardiac-specific promoter. After injection, mESC-derived CMs integrate in the host myocardium and form gap junctions with hosting CMs, which results in significantly improved myocardial contraction in experimental models of heart failure (12). Interestingly, mESC-derived CMs also improved cardiac function in a xenograph sheep model of heart failure (13).
In the late 1990s, human embryonic stem cells (hESCs) came into play. These cells originate from frozen embryos set aside from in vitro fertilization procedures (14). A number of cell lines generated from different institutions throughout the world are today available, with a differentiation potential that varies from cell line to cell line. Kehat et al. (15) provided the first evidence that hESCs can generate differentiated CMs, expressing markers of cardiac differentiation and showing excitation-contraction coupling. The differentiation process is not straightforward: cells are grown on a feeder-layer of fetal fibroblasts, which are usually of mouse origin, while differentiation into EBs, which takes weeks to complete, is induced by seeding small drops of cells on plastic support (15). Within each EB, an area of spontaneous contraction, containing differentiated myocytes, can be more or less extended. The spontaneous depolarization of these cells was further proved by Gepstein’s group (16), who also show that, thanks to their pacemaker property, these cells could be used for therapeutic purposes in a pig model of atrioventricular block. For heart failure treatment, a relatively large number of cells are needed. In this issue of the Journal, Caspi et al. (17) demonstrate that grafting of hESC-derived myocytes improves cardiac function in a rat model of heart failure after MI.
The authors injected undifferentiated hESC, hESC-CMs, or non–hESC-CM derivatives in cyclosporine-immunosuppressed rat myocardium after MI. They found that transplantation of undifferentiated hESCs resulted in the formation of teratoma-like structures, thus indicating that the cardiac environment does not enhance hESC cardiomyogenesis. This phenomenon was prevented by grafting of ex vivo–differentiated hESC-CMs. The grafted CMs, tracked by a combination of elegant cellular and molecular technologies, survived, proliferated, matured, aligned, and formed gap junctions within the transplanted myocardium. Functionally, animals injected with saline or nonmyocyte hESC derivatives demonstrated significant left ventricular dilatation and functional deterioration, whereas grafting of hESC-CMs attenuated the remodeling process. The achieved improvement in cardiac function was dependent on hESC-CMs, demonstrated by deterioration of cardiac function after grafting of non–hESC-CMs. This indicates that the force generated by the transplanted hESC-CM is responsible for functional amelioration, not synthesis of growth factors released by grafted cells. Of note, Caspi et al. (17) found that these cells are devoid of arrhythmogenic potential, a complication that hampers the clinical use of autologous skeletal muscle cell grafting in heart failure (18).
Overall, the current study represents a milestone in this area of investigation. Of course, many issues still need to be resolved before the clinical use of hESC-CMs becomes a reality. First, it is critical to increase the yield of hESC-CMs. Caspi et al. (17) obtained a significant improvement of cardiac function using a relatively small number of cells, considering that cells were selected from the beating part of EBs. This is an encouraging result, but upscaling of the procedure is still a fundamental requirement if large areas of the myocardium have to be savaged. A better knowledge of the biochemistry of the differentiation process of hESC-CM will certainly improve the technical conditions necessary for achieving this task. Secondly, the immunological mismatch between donor and grafted tissues represents another serious limitation. Human endothelial stem cells are less immunogenic that adult cells because they express low levels of the class I major histocompatibility complex (19). One solution to the donor–recipient mismatch problem could be the banking of a large numbers of frozen hESC lines, matching as many potential recipient individuals as possible, an approach similar to the current procedures for bone marrow transplantation. Another solution could be nuclear transfer, a technique in which nuclei from an adult individual, in this case the recipient, substitute oocyte nuclei. Matching zygotes and hESC lines are then developed, a technique named "therapeutic" cloning (10).
The next couple of years will tell us whether hESC-CMs can be considered a realistic therapeutic option in heart failure.
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Footnotes
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* Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology. 
2 Dr. Condorelli is funded by the National Institutes of Health, NHLBI (HL078797-01A1), and by the Italian Ministries of University Research (MUR) and Health.
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References
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