Skip Navigation

European Journal of Heart Failure 2003 5(6):751-757; doi:10.1016/S1388-9842(03)00157-0
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Blatt, A.
Right arrow Articles by Vered, Z.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Blatt, A.
Right arrow Articles by Vered, Z.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© 2003 European Society of Cardiology

Improved regional left ventricular function after successful satellite cell grafting in rabbits with myocardial infarction

Alex Blatta,*, Dror Robinsonb, Gad Cottera, Shai Efratia, Yariv Simantovc, Ilan Bard, Edo Kaluskia, Ricardo Krakovera, Stanislav Sidenkoa, Zoharia Evronb, Lidia Lipab, Natalia Posternakb, Zvi Nevob and Zvi Vereda

a Cardiology Department, Assaf Harofeh Medical Center Zerifin 70300, Israel
b Clinical Biochemistry Department, Sackler School of Medicine Tel Aviv University, Israel
c Animal Experimental Unit, Assaf Harofeh Medical Center Zerifin 70300, Israel
d Thoracic Surgery Department, Assaf Harofeh Medical Center Zerifin 70300, Israel

* Corresponding author. Tel.: +972-8-9779735, fax: +972-8-9228141 E-mail address: alexblatt{at}asaf.health.gov.il


   Abstract
Top
 Notes
 Abstract
1. Introduction
 2. Material and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 
Objective: To evaluate whether satellite cells injected into infarct areas in rabbits remain viable during 6 weeks follow-up and can improve cardiac function as assessed by echocardiography.

Methods: Myocardial infarction was induced in 16 New Zealand white rabbits, by ligation of the marginalis sinistra artery. Tissue from gluteus muscle biopsies was dissected into small pieces and cultured. Within 2–3 weeks the cells were expanded by 2–3 orders of magnitude and were fluorescent labeled. Single cell pellets for resuspension at >106/1 ml were directly injected into the infarct areas in 8 rabbits. In 8 additional rabbits, 1 ml saline was injected (control). Regional left ventricular function was assessed weekly by 2-D echocardiography until animals were sacrificed. Analysis was performed blind and independently by two experienced echocardiographers, based on the American Society of Echocardiography scheme.

Results and discussion: Six treated and five control rabbits completed the study. One week after the artery occlusion, left ventricular function scoring did not differ between groups, mean 8.7±1.6 vs 8.3±1.9 (P=0.74). At 6 weeks post-injection, echocardiographic score was significantly better in the treated group, mean 2.6±0.9 vs 6.9±2.1 (P=0.002). The treated group showed significant gradual segmental improvement between the first week up to week 6. After sacrifice, macro and microscopic transmural areas showed typical changes of myocardial infarction. Histochemical staining identified viable grafted cells in high density 6 weeks post-transplantation in all grafted hearts.

Conclusion: Autologous satellite cells (skeletal myofiber), can be successfully grafted into rabbit hearts following myocardial infarction and may induce improved regional left ventricular function.

Key Words: Cells • Transplantation • Heart failure • Myocardial infarction

Received January 16, 2003; Revised April 21, 2003; Accepted July 31, 2003


   1. Introduction
Top
 Notes
 Abstract
 1. Introduction
2. Material and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 
Congestive heart failure (CHF) is a worldwide growing epidemic, which results in significant disability and mortality in aging populations [1]. Coronary artery disease is the leading etiology. Better treatment of coronary artery disease and acute ischemic syndromes that prevent early mortality following myocardial infarction results in the growing population of survivors with left ventricular dysfunction destined to develop the heart failure syndrome [1]. Heart failure remains a lethal condition, with 5-year survival rates <40% from the time of diagnosis. Since CHF represents the number one public health problem in cardiovascular medicine, its economic impact is tremendous [2].

The cellular basis for the syndrome of heart failure with left ventricular dysfunction is a lack of stem cells in the heart and the inability of the damaged heart cells to undergo repair or divide [3]. Autologous cell grafting by mature or stem lineage expanded in vitro, is now a promising strategy to repair organ damage [4]. Some of these strategies have been demonstrated as efficacious in experimental animal models and may apply to humans [5–7]. One of the most interesting approaches to strengthen the injured or weakened heart, focuses on the strategies to replace dysfunctional, necrotic or apoptopic cardiomyocytes with new cells of mesodermal origin [8,9]. In the previous experimental animal models, cells of various sources have been employed successfully for transplantation; e.g. skeletal myoblast [10,11] fetal cardiomyocytes [12], bone marrow stem cells [13–15]. However, many of the studies had serious drawbacks, such as non-physiologic heart injury, and unreliable methods to follow the left ventricular function [10,11,14,15]. One attractive cell population is the satellite cells, mature muscular skeletal stem cell. The plasticity of skeletal muscle in response to electrical depolarization suggests that individual myoblasts might be converted to muscle fibers capable of performing cardiac work [2].


   2. Material and methods
Top
 Notes
 Abstract
 1. Introduction
 2. Material and methods
3. Results
 4. Discussion
 5. Conclusions
 References
 
2.1. Rabbit cardiac injury model
This study was conducted in accordance with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH publication No. 85-23, revised 1996). Fig. 1 illustrates the study protocol design. Sixteen New Zealand white rabbits, weighing 3.0–4.0 kg, were anesthetized with IV Phenobarbital 0.1 mg/kg. The heart was exposed via mid thoracotomy under strictly sterile conditions in the operating room. Myocardial infarction was induced by ligation of the marginalis sinistra artery immediately after the left main stem bifurcation or trifurcation [16]. Post-operative IM Morphium 0.1 mg/kg was injected three times to relieve myocardial and post-operative pain.


Figure 1
View larger version (32K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 1 Study design: Chronological procedures in all rabbits. The only difference was the sham injection in the control group as opposed to cell grafting in the treated group.

 
2.2. Skeletal myoblast isolation and culture conditions
Muscle biopsies from rabbit gluteus were collected percutaneously by single action biopsy device 16 g (1.6 mm)x15 cm, 20 mm throw, 15 mm notch (US Biopsy, Promex Inc.) under strictly sterile conditions in the operating room. The tissue explants were dissected into small pieces (2 mm3 cubes), which were left to dry by attachment to a vertical standing culture dish, where tissue pieces are held against gravity. Minimal essential medium (MEM) enriched by 10% fetal calf serum, glutamine antibiotics and antimycotics (Beit Ha'amek Industries) were added. After a few days, single cells began to migrate out of the explants forming a monolayer: when the monolayer fans reached significant areas with cell densities close to confluence, the cells were subcultured. Within 2–3 weeks, the cells were expanded by 2–3 orders of magnitude. Cell phenotypes were assessed by immunohistochemistry for myogenin.

2.3. Satellite cell labeling and transplantation
Two days prior to cell collection and implantation, the cells in the monolayers underwent a fluorescent labeling procedure. The cultures were incubated for the final 48 h with a lipophilic, carbocyanine, red fluorescent dye CM-DIL C-7000 chondromethyl benzamide (Molecular Probes, Eugene, Oregon, USA). Final concentration in medium of cell tracer for intracellular cytoplasmatic membranes was 1 mM (dilution 1:1000 in PBS, from stock, 50 µg of dye in 50 µl of DMSO (dimethylsulfoxide)). After the incubation period in the presence of the fluorescent dye, the cultures were washed thoroughly (three times) by PBS, prior to trypsinization. Single cell pellets for resuspension at >106/1 ml were intramurally implanted by direct injection into the infarct areas in eight rabbits. In the other eight rabbits, 1 ml saline was injected without cells (control). Following killing, the hearts were dissected and inspected by fluorescent microscope. The phenotypic fate (differentiation status) of the injected cells concentrated in the ischemic regions, was followed using cardiomyocyte specific markers. Pancadherin, desmine and myogenine were positive, while CD-34 and connexin 43 were negative.

2.4. Clinical follow-up
Animals were evaluated by ECG and two-dimensional echocardiography (Hewlett Packard, Sonos 1000) before and after each procedure and on a weekly basis until the animals were killed. In each animal parasternal, long and short axis views were obtained and recorded on 0.5-inch video cassettes (VHS) for off-line analysis. An attempt was made to best display the damaged area, Two-dimensionally guided M-mode tracings were also obtained at the damaged area. Regional left-ventricular function was assessed by two experienced echocardiographers independently, both blinded to the data, based on the recommendations of the American Society of Echocardiography scoring scheme [17]. The result of wall motion score throughout the experiment from both reviewers were averaged and plotted.


   3. Results
Top
 Notes
 Abstract
 1. Introduction
 2. Material and methods
 3. Results
4. Discussion
 5. Conclusions
 References
 
3.1. Myocardial infarction documentation
Six rabbits in the experimental group and five in the control group completed the entire study. Five animals died a few hours post operation. There were no further deaths in the two groups during the 6 weeks following the induction of the experimental myocardial infarction. ECG and echocardiography confirmed the presence of myocardial infarction in all animals. Main artery ligation lead to transmural wall necrosis, in all the adult rabbit hearts as shown by enzymatic dye (Fig. 2a). The infarct area was between 25 and 40% of the left ventricular mass. Histological changes of the acute myocardial infarction in the rabbits were similar to those seen in humans by light microscopy (Fig. 2b and c).


Figure 2
View larger version (41K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 2 Pathological transmural myocardial infarction identification. (a) In this heart cross-section cut a transmural wall myocardial necrosis is detected by viable staining. (b and c) Panoramic microscopic section of heart muscle shows the transmural involvement, and in the magnified section, the typical histological and cytological myocardial infarction changes: edema, waviness of fibers and piknotic nucleus.

 
3.2. Echocardiographic wall motion score
The mean echocardiographic wall motion score was similar in the two groups at the time of the injection, 8.7±1.6 in the experimental group vs. 8.3±1.9 in the control group (P=0.74). One week after the injection there was a trend towards worsening in wall motion score in the experimental group with score of 9.25±0.9, as compared with 7.8±2.1 in the control group (P=0.16). From the second week onwards, wall motion score in the experimental group improved progressively, reaching significance, starting from the third week. Six weeks after the injection the echocardiographic wall motion score was 2.6±0.9 in the experimental group as compared with 6.9±2.1 in the control group (P=0.002). The weekly echocardiography wall motion scores are summarized in Table 1 and Fig. 3. Fig. 4 shows one of the grafted rabbits at baseline, following induction of myocardial infarction and 6 weeks following cell transplantation. The treated group showed better segmental improvement between the first week to the end of the follow-up, than the control group, mean improvement of 6.1±1.8 vs. 1.4±0.6 (P<0.0001).


View this table:
[in this window]
[in a new window]

  		Table 1 Echocardiographic wall motion scoring

 


Figure 3
View larger version (9K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 3 Mean echocardiographic wall motion score during the follow-up period. The mean echocardiographic wall motion score was similar in the two groups at the time of the injection, 8.7 in the study group vs. 8.3 in the control. Six weeks after injection, the echocardiographic wall motion score was 2.6 in the study group as compared with 6.9 in the control group (P=0.002).

 


Figure 4
View larger version (87K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 4 Two-dimensionally guided M-mode echocardiography obtained from rabbit number 8 at baseline (panel A) showing normal septal function; at 24 h post-induced infarction (coronary artery ligation) and satellite cell transplantation (panel B) showing an almost akinetic septum; and at 6 weeks post-transplantation (panel C) demonstrating full recovery of septal motion and thickening.

 
3.3. Post-euthanasia identification of cell grafting
Histochemical staining identified viable grafted cells in the damaged area in high density, 6 weeks post-transplantation in all transplanted hearts (Fig. 5). The transplanted cell distribution was around the needle injection site within the necrotic area borders in most of the cases and extended beyond them in some cases. No evidence for cell rejection at 6 weeks was shown. Autofluorecence was eliminated by calibrating the system with the control group. In the cell culture, prior to transplantation, >95% of the cells yielded positive staining for desmin indicating that indeed the antibodies utilized recognize desmin (Fig. 6).


Figure 5
View larger version (61K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 5 Viable labeled myoblast graft identification. (a) In vitro aspect of labeled smear of expanded satellite muscle cells before transplantation. (b) Histochemical staining identified viable grafted cells in high density 6 weeks post-transplantation in all transplanted hearts by confocal microscopy, panoramic (b) and magnified views (c).

 


Figure 6
View larger version (105K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 6 Cultures of satellite cells derived from autologous muscle confirmed by staining with antidesmine antibodies. Other cells show negative staining. Photomicrograph x100 (a) and x40 (b).

 

   4. Discussion
Top
 Notes
 Abstract
 1. Introduction
 2. Material and methods
 3. Results
 4. Discussion
5. Conclusions
 References
 
Myoblasts have already been successfully grafted into injured hearts in many experimental animal models [8–12,14,15] and recently in the phase I clinical trial [18]. The current study illustrates certain unique aspects: Firstly, previous studies have used non-physiologic myocardial tissue damage such as the cryoinjury [10] or transient coronary ligation [8,9]. The latter technique produces an occlusion/reperfusion model with a large area of stunned myocardium. The current study utilizes a model that mimics very closely acute myocardial infarction without reperfusion, a permanent occlusion of a major coronary artery. Indeed, the damage, as documented on ECG and echo, and subsequently on pathologic specimens was mostly transmural (Fig. 2). Secondly, we chose to inject satellite cells, as opposed to embryonic cells or an alternative source such as: adult skeletal myoblast; fetal cardiomyocytes; bone marrow stem cells; and multipotential mesenchymal progenitor stem cells [19]. We believe that satellite cells may offer a valid and practical approach, and the subsequent studies should confirm the current results. Moreover, they can replicate and augment exponentially in vitro to achieve bigger inoculum transplantation initiated from a small tissue biopsy. Our data indicate that these cells could be easily injected into the damaged areas and that they remain viable up to 6-weeks post-transplantation. Based on the new concept of cells phenotype plasticity, the cultured satellite cells yielding a homotypic cell culture, where almost every cell (above 95%) yielded positive staining, employing the antibodies against desmine. Desmine serves as a marker for satellite progenitor stem cells. These data confirm the viability of satellite cells in infarct tissue at least for 6-weeks. The fetal source has ethical and quantity limitations: they do not replicate or perhaps only once in vitro, which implies the necessity of multiple fetal donors to achieve the transplantation sample. Another promising horizon is the satellite cell modification genetically by transfection [20–22] to highlight contractile and electrical properties that allow enhancement of cardiac work. The current study was not designed to investigate the exact morphologic changes these cells undergo following transplantations. Nevertheless, our data indicate that, based on a carefully designed echo scoring method [17], which has not yet been applied in other experiments, it would seem that wall motion improves after cell transplantation compared with the placebo group. Regional wall motion score analysis on two-dimensional echocardiography is a well-accepted technique to evaluate and follow-up left ventricular function. In the current study, two individuals assessed wall motion scores, both experienced in echocardiography, each analyzing the data in a blind fashion, and eventually their scores were averaged to provide the final results. Both reviewers’ results were very close, although one reviewer tended to assign more significant wall motion scores in a consistent way throughout the analysis. Our echocardiographic data concords with the results recently published by Jain et al. [11], who also reported beneficial effects of cell therapy 6-weeks after transplantation.

Several hypotheses can explain this improvement. By successfully grafting into the damaged zone, we may induce transformation of a transmural into a non-transmural myocardial infarction. This may be accompanied by additional benefit: decrease in the thinning and dilatation process; and expansion [3,23,24], thereby improving remodeling of the injured heart and geometry preservation. Secondly, if we assume the presence of mechanical or electrical coupling between the grafted cells and host myocardium, as described by Connold et al. [25] and the ability of skeletal muscle cells to transform into indefatigable muscle, that is, expression of pre-dominant slow-twitch isoforms of contractile and sarcoplasmic reticulum proteins investigated by Jolez et al. [26], the high grade transplanted cells in the injured and debilitated areas contribute to myocardial performance. The possibility of transdifferentiation [3,27] phenomenons by the myoblast can contribute to this electromechanical coupling. Thirdly, if this coupling and changes do not occur, or the grafted cells were encased in scar, the presence of the implanted myoblasts exert favorable effects by changing the scare quality and properties with beneficial passive mechanical effect [3,24].

There may be many limitations in the current approach. These include possible tumor formation, genetic drift of the cell line or electrical interference [20]. The first two points can be attenuated by autologous cell graft strategy, that minimizes the risk of neoplasia and immune rejection and prevent immunosuppression therapy associated with allogenic or xenogenic cells source with respect to electrical instability. Electrical interference and the development of an arrhythmogenic focus is a theoretically possible negative side effect of this strategy. At 6 weeks, our findings showed lack of integration between the transplanted cells and host myocardium, indicating impaired electrical coupling. This intramyocardial heterogeneous focus, which includes exitable cells, may theoretically become arrhythmogenic. Alternatively, re-entry pathways created by cell grafting could increase the arrhythmogenic possibility. Menasche et al. [18] in the first phase I clinical trial, implanted internal automatic defibrillators in four of 10 grafted patients due to episodes of sustained ventricular tachycardia, which in only one case were symptomatic. In our animal study, none of the transplanted rabbits showed late sudden death. Yet, no doubt, further studies will be required to elucidate these important issues.


   5. Conclusions
Top
 Notes
 Abstract
 1. Introduction
 2. Material and methods
 3. Results
 4. Discussion
 5. Conclusions
References
 
We believe that the current experiment allows us to conclude that satellite cells and skeletal myofiber, can be successfully grafted into rabbit hearts following myocardial infarction and may induce improved regional left ventricular function. These experiments carried out in rabbits may apply to humans, and perhaps become clinically important.


   Notes
Top
 Notes
Abstract
 1. Introduction
 2. Material and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 
{star} Presented in part in the ‘Young Investigator Award Competition’, Basic Science category, European Society of Cardiology Congress, Stockholm 2001.


   References
Top
 Notes
 Abstract
 1. Introduction
 2. Material and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 

  1. Starling R.C. The health-care impact of heart failure. In: Textbook of cardiovascular medicine—Topol E.J., ed. (1998) Philadelphia, PA: Lippincott-Raven Publishers. 2205–2214.
  2. Kannel W.B., Ho K., Thom T. Changing epidemiological features of cardiac failure. Br Heart J (1994) 72(Suppl):S3–S9.[Free Full Text]
  3. Kessler P.D., Byrne B.J. Myoblast cell grafting into heart muscle: cellular biology and potential applications. Annu Rev Physiol (1999) 61:219–242.[CrossRef][Web of Science][Medline]
  4. Boheler K.R., Fiszman M.Y. Can exogenous stem cells be used in transplantation? Cells Tissues Organs (1999) 165:237–245.[CrossRef][Web of Science][Medline]
  5. Tsai R.J.T., Li L.M., Chen J.K. Reconstruction of damaged corneas by transplantation of autologous limbal epithelial cells. N Engl J Med (2000) 343:86–93.[Abstract/Free Full Text]
  6. Freed C.R., et al. Transplantation of embryonic dopamine neurons for severe Parkinson's disease. N Engl J Med (2001) 344:710–719.[Abstract/Free Full Text]
  7. Nevo Z., Robinson D., Horowitz S., et al. The manipulated mesenchymal stem cells in regenerated skeletal tissues. Cell Transplant (1998) 7:63–70.[CrossRef][Web of Science][Medline]
  8. Murry C.H., Wieseman R.W., Schwartz S.M., et al. Skeletal myoblast transplantation for repair of myocardial necrosis. J Clin Invest (1996) 98:2512–2523.[Web of Science][Medline]
  9. Leor J., Patterson M., Quinones M.J., et al. Transplantation of fetal myocardial tissue unto the infracted myocardium of rat. Circulation (1996) 94(Suppl_II):II-332–II-336.
  10. Taylor D.A., Atkins B.Z., Hungspreugs P., et al. Regenerating functional myocardium: Improve performance after skeletal myoblast transplantation. Natl Med (1998) 4:929–933.[CrossRef]
  11. Jain M., DerSimonian H., Brenner D.A., et al. Cell therapy attenuates deleterious ventricular remodeling and improves cardiac performance after myocardial infarction. Circulation (2001) 103:1920–1929.[Abstract/Free Full Text]
  12. Reinecke H., Zhang M.Z., Bartosek T., et al. Survival, integration, and differentiation of cardiomyocyte graft. Circulation (1999) 100:193–202.[Abstract/Free Full Text]
  13. Tomita S., Li R.K., Weisel R.D., et al. Autologous transplantation of bone marrow cells improves damaged heart function. Circulation (1999) 100(Suppl II):II247–II256.[Medline]
  14. Orlic D., Kajstura J., Chimenti S., et al. Bone marrow cells regenerate myocardium. Nature (2001) 410:701–705.[CrossRef][Medline]
  15. Ferrari G., Cusella-Deangelis G., Coletta M., et al. Muscle regeneration by bone marrow-derived myogenic progenitors. Science (1998) 279:1528–1530.[Abstract/Free Full Text]
  16. Podesser B., Wollenek G., Seitelberger R., et al. Epicardial branches of the coronary Arteries and their distribution in the rabbit heart: the rabbit heart as a model of regional ischemia. Anat Rec (1997) 247:521–527.[CrossRef][Medline]
  17. Bourdillon P.V.D., Broderick T.M., Sawada S.G., et al. Regional wall motion index for infracted and non-infracted regions after reperfusion in acute myocardial infarction: comparison with global wall motion index. J Am Soc Echocardiogr (1989) 2:398–402.[Medline]
  18. Menasche P., Hagege A.A., Vilquin J.T., et al. Autologous skeletal myoblast transplantation for severe post-infarction left ventricular dysfunction. J Am Coll Cardiol (2003) 41:1078–1083.[Abstract/Free Full Text]
  19. Liechty K.W., MacKenzie T.C., Shaaban A.F., et al. Human mesenchymal stem cells engraft and demonstrate site-specific differentiation after in utero transplantation in sheep. Natl Med (2000) 6:1282–1286.[CrossRef]
  20. Leor J., Prentice H., Sartorelli V., et al. Gene transfer and cell transplant an experimental approach to repair a ‘broken heart’. Cardiovasc Res (1997) 35:431–441.[Free Full Text]
  21. Blau H.M., Springer M.L. Muscle-mediated gene therapy. N Engl J Med (1995) 333:1554–1556.[Free Full Text]
  22. Yla-Herttuala S., Martin J.F. Cardiovascular gene therapy. Lancet (2000) 355:213–222.[CrossRef][Web of Science][Medline]
  23. Yun Y.L., Li Y.Y., McTiernan Ch.F., et al. Interplay of matrix metalloproteinases, tissue inhibitors of metalloproteinases and their regulators in cardiac matrix remodeling. Cardiovasc Res (2000) 46:214–215.[Abstract/Free Full Text]
  24. Etzion S., Battler A., Barbash I., et al. Influence of embryonic cardiomyocyte transplantation on the progression of heart failure in a rat model of extensive myocardial infarction. J Mol Cell Cardiol (2001) 33:1321–1330.[CrossRef][Web of Science][Medline]
  25. Connold A.L., Frischknecht R., Dimitrakos M., et al. The survival of embryonic cardiomyocytes transplanted into damaged myocardium. J Muscle Res cell Motil (1997) 18:63–70.[CrossRef][Web of Science][Medline]
  26. Jolesz F., Sreter F.A. Development, innervation and activity pattern-induced changes in skeletal muscle. Annu Rev Physiol (1981) 43:531–532.[CrossRef][Web of Science][Medline]
  27. Anderson D.J., Gage F.H., Weissman I.L. Can stem cells lineage boundaries? Natl Med (2001) 7:393–395.[CrossRef]

Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?



This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Blatt, A.
Right arrow Articles by Vered, Z.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Blatt, A.
Right arrow Articles by Vered, Z.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?