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Cardiovascular Research 2003 58(2):241-245; doi:10.1016/S0008-6363(03)00317-1
© 2003 by European Society of Cardiology
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Copyright © 2003, European Society of Cardiology

Spotlight on stem cells—makes old hearts fresh

Marcel A.G. van der Heydena,*, Jürgen Heschelerb and Christine L. Mummeryc

aDepartment of Medical Physiology, University Medical Center Utrecht, P.O. Box 85060, 3508 AB Utrecht, The Netherlands
bDepartment of Neurophysiology, University of Cologne, Robert-Koch-Str. 39, 50931 Köln, Germany
cHubrecht Laboratory, Netherlands Institute of Developmental Biology, Uppsalalaan 8, 3584 CT, The Netherlands

m.a.g.vanderheyden{at}med.uu.nl

* Corresponding author. Tel.: +31-30-253-8418; fax: +31-30-253-9036.

Received 21 February 2003; accepted 22 February 2003

...makes old hearts fresh... A winter's tale; William Shakespeare


   1 Historical perspective
Top
 1 Historical perspective
2 Introduction to the...
 3 Future perspectives
 Acknowledgments
 References
 
Is there a better way to restore function of a damaged organ than simply by replacing it?

The idea of transplantation has caught the imagination throughout history, leading to the creation of mythological figures like the sphinx, mermaids and centaurs; all examples of xenotransplantation. Centuries later, transplantation fantasy and technology came into the hands of the medical profession, with Mary Shelley's Frankenstein (1818) being an extreme example from classical literature, which, fortunately, does not reflect clinical results of those times.

Modern stem cell based transplantation in humans originates in blood and bone marrow research. In the late 19th century, the concept of blood forming stem cells was formed [1–3]. This important concept was further studied and elaborated in the next decades by, among others, Pappenheim and Ferrata. The first therapeutic use of bone marrow was described in 1896. Here, bone marrow extract was given orally as treatment for leukemia [4]. Bone marrow cell transplantation was first performed by Schretzenmayr in 1937 when he injected freshly aspirated bone marrow intramuscularly to treat parasitic infection [5]. Soon afterwards, in 1939, Osgood infused bone marrow intravenously to correct primary bone marrow disorders [6]. Since then, interest in bone marrow transplantation research and therapy continued to increase, especially during the Cold War as a potential therapy for otherwise lethal irradiation damage. During the late 1960s successful bone marrow transplantations were reported in man [7,8]. Today, bone marrow transplantation is a routine treatment for various haematopoietic diseases.

While research on adult stem cells originates in blood and bone marrow research, that on embryonic stem cells derives from studies on fertility and a particular tumor known as teratocarcinoma. These tumors consist of many differentiated tissues but contain a subpopulation with an undifferentiated phenotype. Tumors lacking these undifferentiated embryonal carcinoma stem cells are benign, while those containing stem cells niches continue to grow and become invasive [9]. Understanding stem cell differentiation was thus thought to hold the key to controlling malignancy. Deriving embryonal carcinoma cell lines in culture was an important step forward, and was first described in 1967 [10]. Since then, numerous cell lines have been derived from human and murine teratocarcinomas. Differentiation of these cells could be initiated by cell aggregation resulting in embryo-like structures called embryoid bodies, which were initially observed in ascites from mice containing embryonal carcinomas (reviewed in [11]). It turned out that these cells were a valuable model system for developmental biologists too, since their differentiation pattern in vitro recapitulated aspects of embryogenesis and resulted in the formation of cells representative of all three germ layers.

During the same period, attempts to derive human embryonic stem cells were initiated using surplus embryos from in vitro fertilization clinics. These were initially unsuccessful but in an attempt to study the differentiation of isolated cells from preimplantation blastocysts, rabbit embryonic cells were brought into culture [12]. These embryonic stem cells (or ES cells) were able to form chimaeric embryos proving their pluripotency but also represented an important technical advance, later widely used in the mouse [13]. The resulting knock-out as well as knock-in (transgenic) mice opened a wide new horizon of understanding genetic diseases and functional genomics.

In 1998, more than a decade after first attempts, the first human ES cells were isolated into culture [14]. A month later, embryonic germ cells were derived from human primordial germ cells [15]. Almost as an ethical reaction against these cells, research on adult stem cells was intensified and a remarkable plasticity of these cells lead to publications with titles like ‘muscle from bone marrow’, ‘turning brain into blood’ and ‘turning blood into brain’ (i.e. [16–18]). Since these first studies on stem cells, a plethora of articles has appeared on both adult and embryonic stem cells, all with the same purpose in mind: stem cells based therapy. Whether this is all hype and hope is not yet clear. However, the interest has stimulated the editors of Cardiovascular Research to dedicate this entire issue to stem cells and their potential applications in the cardiovascular field. This issue thus contains thirteen review articles and ten state-of-the art original papers on stem cell biology. Table 1 classifies the contributions into four main themes, which are discussed in more detail below.


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  		Table 1 Thematic relationships among papers

 

   2 Introduction to the reviews and original contributions
Top
 1 Historical perspective
 2 Introduction to the...
3 Future perspectives
 Acknowledgments
 References
 
2.1 Embryonic development and phenotyping
Embryos are capable of generating a fully functional heart from primitive precursors. The study of heart development can thus provide vital clues on how to induce stem cells to differentiate into heart and vessel cells. Therefore, three reviews cover heart development and morphogenesis. Solloway and Harvey focus on the activation of signal transduction pathways during the later phases of heart development [19]. Habets et al. describe the activity of a number of cardiac specific promoter–reporters in vivo and thereby provide positional information on signaling events during heart morphogenesis [20]. Fijnvandraat et al. provide an overview of cardiac specific markers for different regions of the heart and conclude that marker expression in adult hearts is more restricted than in embryonic hearts and hence the use of current markers is questioned [21]. Luttun and Carmeliet describe the development of the cardiac vasculature and the potential of vascular stem cells for specific therapeutic purposes [22].

It has been proposed that stem cells may be a source of myocytes for cell replacement therapy in defective adult hearts. It could be argued that donor cells resembling adult myocytes would be the best possible candidates. This comparison, however, requires a full characterization of adult cardiomyocytes of different origins within the human heart. Bird et al. describe human adult atrial and ventricular cardiomyocytes in culture and characterized these at the mRNA and protein level for a variety of markers [23]. Isolated human cardiac myocytes as described may be an important model for studying interaction and communication between primary adult and human stem cell-derived cardiomyocytes.

2.2 Stem cell sources and models, differentiation and characterization
It is still a matter of debate whether and which source of stem cells will be useful for cell transplantation in the clinic. It may even turn out that each type of therapy may have a favored stem cell source. Passier and Mummery review the relative merits of embryonic versus adult cells for possible use in a variety of diseases [24]. Several other reviews focus on specific stem cell types, their characteristics and use for cardiac research. P19 embryonal carcinoma (EC) cells have been available for 21 years, but are still a favored model system for studying cardiomyocyte differentiation and physiology as reviewed by Van der Heyden and Defize [25]. Mouse ES cells are another stem cell model that contributed to our understanding of cardiac differentiation and stem cell derived cardiomyocyte electrophysiology, as shown by Sachinidis et al. [26]. These two stem cell models have paved the way for the more recently established human ES cells; rapid developments of the last four years are reviewed by Gerecht et al. [27]. The isolation of endothelial progenitor cells (EPCs) was a key step in the concept of therapeutic vasculogenesis. The review of Masuda and Asahara highlights an update of EPC biology and the potential for therapeutic regeneration [28].

Electrophysiological data of P19 EC cell derived cardiomyocytes were rather limited in literature. The original article of Van der Heyden et al. describes the establishment of the electrophysiological system during cardiac differentiation of these cells, using a combination of molecular and electrophysiological techniques [29]. A similar combination of techniques was used by Fijnvandraat et al. to establish that mES derived cardiomyocytes resemble embryonic cardiomyocytes from the primary heart tube [30]. It was demonstrated earlier that mouse bone marrow stromal cells treated with 5'-azacytidine differentiate into cardiomyocytes [31]. Liu et al. seriously question whether the same phenomenon occurs in rat derived BMSCs [32]. Recently, it was established that stem cells could be effectively recovered from cryopreserved human cord blood after a 15-year storage period [33]. In this issue, Eggermann et al. compared in vitro expansion of human umbilical cord blood derived EPCs either from nonselected or CD34+ preselected mononuclear cells with remarkable results [34].

2.3 Stem cell transplantation
Many studies have already described transplantation of cells into the hearts of animals. The studies have differed in the choice of donor cells, i.e. fetal or neonatal cardiomyocytes, bone marrow derived stem cells and even differentiated mES cells. Often the heart is damaged, for instance by cryoinjury or coronary ligation. Nevertheless, clinical trials are already being performed in humans using skeletal muscle satellite cells as donor cells for repairing hearts with myocardial infarctions. This rapidly expanding literature is reviewed in four excellent papers from Dowell et al., Reffelmann and Kloner, Goldenthal and Marín-García, and Menasché [35–38], each with its own focus.

The original contribution of Johkura et al. uses mES derived cardiomyocytes to generate ectopic or extracardiac ‘hearts’ surrounding large blood vessels [39]. They observed continuous differentiation of the transplant, which became vascularized, perfused by the host and survived for at least 30 days. Agbulut et al. established that bone marrow derived cells can contribute to myocardial repair following acute nonischemic cardiomyopathy. Precursors could be found in unpurified bone marrow but not in Sca-1+ haematopoietic progenitor cell populations [40]. In a technical paper, Chazaud et al. assessed the feasibility of closed chest skeletal myoblast transplantation in pigs using a specialized catheter approach [41]. Finally, two papers demonstrated the potential of EPCs to repopulate the vessel wall. In the case-report of Suzuki et al., a patient suffering from acute radiation syndrome was treated with a gender-mismatched bone marrow transplant. It was established that 25% of all aortic endothelial cells became replaced by donor-origin endothelial cells [42]. In the paper of Griese et al., purified EPCs were modified to express anticoagulants before transplantation. Following balloon injury, engineered EPCs covered 71% of the damaged vessels while sustained secretion of the anticoagulants was detected [43].


   3 Future perspectives
Top
 1 Historical perspective
 2 Introduction to the...
 3 Future perspectives
Acknowledgments
 References
 
As evidenced by the reviews in this special issue, many new insights have been made in the field of cardiovascular research with respect to stem cell biology, differentiation and transplantation. During the last few years, the perspective of clinical application, mainly fed by the discovery of adult stem cell plasticity and generation of human ES cells, has attracted many new researchers to the field. Though progress may seem rapid, many important issues still have to be addressed before stem cell based tissue repair fulfils its promise.

Besides the usage for transplantation, stem cells will also stipulate our future basic knowledge of the cardiovascular system, including developmental physiology and functional genetics. Furthermore, human stem cell derived cardiomyocytes will allow the development of high throughput screening systems enabling the fast determination of pharmacological actions of new chemicals.

For each therapeutic intervention the best possible stem cell has to be used. It may very well be that for one application adult stem cells will prove to be the best, for instance EPCs for vessel regeneration, while ES cells may be the choice for others, for instance a tissue based pacemaker consisting of nodal-like cardiomyocytes or a contracting tissue based on ventricular-like cardiomyocytes. Much research is still required to determine the potential of each different stem cell type.

Currently, none of the existing human ES cell lines can be used for clinical applications since they were all derived and cultured under non-GMP conditions. Therefore, new lines will have to be derived under GMP conditions, but complications such as obtaining inner cell masses from human embryos without the use of animal complement and culturing of the cells in serum and feeder-cell free conditions will have to be solved.

Another problem that has to be tackled is immunological rejection following ES derived cell transplantation. This could be solved by matching and immunosuppressive drugs as used now for organ transplants or by modifying the immunological marking system of the cells themselves. This would require genetic manipulation of the stem cells raising new ethical problems. Fewer immunological problems would be expected if adult stem cells were used for autologous transplantation. The current trials on skeletal myoblast satellite cell transplantation are an example of this already.

When simply transplanted into an individual, undifferentiated ES cells may form teratomas. So it will be essential to ensure no residual stem cells are present in transplants. Another option is to induce differentiation of the cells into the desired phenotype and isolation of these to homogeneity. Other options are irradiation or expression of suicide genes, which may be activated if cells become transformed.

Eventually, every transplantation technology will have to be evaluated extensively in double blind animal models as well as in clinical trials. This also requires clearly defined endpoints to measure the effects of the therapy.

In conclusion, there may eventually be better alternatives for whole organ transplantation, but only when stem cells can be controlled. If sufficiently large donor cell numbers can be generated, this may ultimately lead to stem cell based therapeutic application for cardiovascular repair.

Time for primary review 1 day.


   Acknowledgments
Top
 1 Historical perspective
 2 Introduction to the...
 3 Future perspectives
 Acknowledgments
References
 
The authors thank the members of the previous editorial team of Cardiovascular Research, Tobias Opthof and Allard van der Wal, assigned to this Spotlight issue for their support and help to successfully complete this project. We thank all personnel in the editorial offices in Amsterdam and Giessen for their help in handling the manuscripts efficiently. This editorial was supported by the Netherlands Science Organization, section Medical Sciences (NWO-MW) grant no. 902-16-193 (MvdH).


   References
Top
 1 Historical perspective
 2 Introduction to the...
 3 Future perspectives
 Acknowledgments
 References
 

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