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Cardiopoietic programming of embryonic stem cells for tumor-free heart repair.

Behfar A, Perez-Terzic C, Faustino RS, Arrell DK, Hodgson DM, Yamada S, Puceat M, Niederländer N, Alekseev AE, Zingman LV, Terzic A - J. Exp. Med. (2007)

Bottom Line: Here, the tumorigenic threat associated with embryonic stem cell transplantation was suppressed by cardiac-restricted transgenic expression of the reprogramming cytokine TNF-alpha, enhancing the cardiogenic competence of recipient heart.Characterized by a down-regulation of oncogenic markers, up-regulation, and nuclear translocation of cardiac transcription factors, this predetermined population yielded functional cardiomyocyte progeny.Thus, cardiopoietic programming establishes a strategy to hone stem cell pluripotency, offering a tumor-resistant approach for regeneration.

View Article: PubMed Central - PubMed

Affiliation: Marriott Heart Disease Research Program, Division of Cardiovascular Diseases, Department of Medicine, Mayo Clinic, Rochester, MN 55905, USA.

ABSTRACT
Embryonic stem cells have the distinct potential for tissue regeneration, including cardiac repair. Their propensity for multilineage differentiation carries, however, the liability of neoplastic growth, impeding therapeutic application. Here, the tumorigenic threat associated with embryonic stem cell transplantation was suppressed by cardiac-restricted transgenic expression of the reprogramming cytokine TNF-alpha, enhancing the cardiogenic competence of recipient heart. The in vivo aptitude of TNF-alpha to promote cardiac differentiation was recapitulated in embryoid bodies in vitro. The procardiogenic action required an intact endoderm and was mediated by secreted cardio-inductive signals. Resolved TNF-alpha-induced endoderm-derived factors, combined in a cocktail, secured guided differentiation of embryonic stem cells in monolayers produce cardiac progenitors termed cardiopoietic cells. Characterized by a down-regulation of oncogenic markers, up-regulation, and nuclear translocation of cardiac transcription factors, this predetermined population yielded functional cardiomyocyte progeny. Recruited cardiopoietic cells delivered in infarcted hearts generated cardiomyocytes that proliferated into scar tissue, integrating with host myocardium for tumor-free repair. Thus, cardiopoietic programming establishes a strategy to hone stem cell pluripotency, offering a tumor-resistant approach for regeneration.

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Neoplastic risk of embryonic stem cell therapy averted with transgenic cytokine overexpression. (A) Host heart (WT) demonstrated a finite capacity to secure tumor-free stem cell transplantation, i.e., 100% tumor-free outcome at 3 × 105 cells/heart versus 82% and 32% tumor-free outcomes at 106 cells/heart and 3 × 106 cells/heart, respectively (blue). Transgenic cardiac-restricted overexpression of TNF-α (TNF-TG) increased the capacity of host heart to guide stem cell cardiogenesis ifying uncontrolled growth, i.e., 100% tumor-free outcome at 3 × 106 cells/heart (red). Conversely, deletion of the kinase domain of the TGF-β receptor (ΔTGFβRII) or overexpression of the BMP inhibitor Noggin, both disrupting cardiogenic signaling in embryonic stem cells (ES), precipitated tumorigenesis after transplantation, i.e., 0% tumor-free outcomes at 3 × 105 cells/heart (light and dark green). For each condition, n indicates the number of treated animals. (B and C) TNF-α priming achieved tumor-free incorporation of transplanted stem cells (3 × 106 cells/heart; top panel), with embryonic stem cell–derived cardiomyocytes integrating with host myocardium as determined by CFP expression (B, bottom and C). Bars: (B, top) 2 mm; (B, bottom and C) 10 μm. (D) Transgenic TNF-α overexpression (TNFα-TG) up-regulated myocardial p38 MAPK and TGF-β expression with α-actinin used as a control for protein loading. (E and F) Blockade of the TGF-β superfamily through overexpression of Noggin or ΔTGFβRII precipitated tumor formation because of uncontrolled growth of implanted embryonic stem cells even at 3 × 105 cells/heart.
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fig2: Neoplastic risk of embryonic stem cell therapy averted with transgenic cytokine overexpression. (A) Host heart (WT) demonstrated a finite capacity to secure tumor-free stem cell transplantation, i.e., 100% tumor-free outcome at 3 × 105 cells/heart versus 82% and 32% tumor-free outcomes at 106 cells/heart and 3 × 106 cells/heart, respectively (blue). Transgenic cardiac-restricted overexpression of TNF-α (TNF-TG) increased the capacity of host heart to guide stem cell cardiogenesis ifying uncontrolled growth, i.e., 100% tumor-free outcome at 3 × 106 cells/heart (red). Conversely, deletion of the kinase domain of the TGF-β receptor (ΔTGFβRII) or overexpression of the BMP inhibitor Noggin, both disrupting cardiogenic signaling in embryonic stem cells (ES), precipitated tumorigenesis after transplantation, i.e., 0% tumor-free outcomes at 3 × 105 cells/heart (light and dark green). For each condition, n indicates the number of treated animals. (B and C) TNF-α priming achieved tumor-free incorporation of transplanted stem cells (3 × 106 cells/heart; top panel), with embryonic stem cell–derived cardiomyocytes integrating with host myocardium as determined by CFP expression (B, bottom and C). Bars: (B, top) 2 mm; (B, bottom and C) 10 μm. (D) Transgenic TNF-α overexpression (TNFα-TG) up-regulated myocardial p38 MAPK and TGF-β expression with α-actinin used as a control for protein loading. (E and F) Blockade of the TGF-β superfamily through overexpression of Noggin or ΔTGFβRII precipitated tumor formation because of uncontrolled growth of implanted embryonic stem cells even at 3 × 105 cells/heart.

Mentions: The propensity for tumorigenic outcome correlated with embryonic stem cell load (Fig. 2 A). The neoplastic threat associated with stem cell transplantation was eliminated by cardiac-restricted transgenic expression of the stress cytokine TNF-α (TNF-α–TG), averting tumorigenic outcome even at doses of 10,000 cells per mg of myocardial tissue (∼3 × 106 stem cells/heart) that produced uncontrolled growth in wild-type hearts (Fig. 2 A). Up-regulated myocardial expression of TNF-α secured cardiac differentiation of implanted embryonic stem cells with proper integration within host myocardium occurring over an increasing range of stem cell loads (Fig. 2, A and B). Autofluorescence and cell fusion was ruled out by multiwavelength confocal visualization after immunohistochemical and nuclear probing (Fig. 2 C). The in vivo action of TNF-α was associated with myocardial up-regulation of the p38 mitogen-activated protein kinase and enhanced expression of the cardiogenic growth factor TGF-β (Fig. 2 D). Deletion of the TGF-β receptor kinase domain (ΔTGF-βRII) in embryonic stem cells or disruption of the stem cell ability to recognize members of the TGF-β superfamily, i.e., BMP through overexpression of the BMP inhibitor, noggin, prevented cardiac differentiation leading to tumor formation even at low stem cell loads (Fig. 2 A, E, and F). Thus, the tumorigenic risk of embryonic stem cells can be blunted by TNF-α cardiac-restricted overexpression or exaggerated by removal of the TGF-β superfamily guidance of transplanted cells.


Cardiopoietic programming of embryonic stem cells for tumor-free heart repair.

Behfar A, Perez-Terzic C, Faustino RS, Arrell DK, Hodgson DM, Yamada S, Puceat M, Niederländer N, Alekseev AE, Zingman LV, Terzic A - J. Exp. Med. (2007)

Neoplastic risk of embryonic stem cell therapy averted with transgenic cytokine overexpression. (A) Host heart (WT) demonstrated a finite capacity to secure tumor-free stem cell transplantation, i.e., 100% tumor-free outcome at 3 × 105 cells/heart versus 82% and 32% tumor-free outcomes at 106 cells/heart and 3 × 106 cells/heart, respectively (blue). Transgenic cardiac-restricted overexpression of TNF-α (TNF-TG) increased the capacity of host heart to guide stem cell cardiogenesis ifying uncontrolled growth, i.e., 100% tumor-free outcome at 3 × 106 cells/heart (red). Conversely, deletion of the kinase domain of the TGF-β receptor (ΔTGFβRII) or overexpression of the BMP inhibitor Noggin, both disrupting cardiogenic signaling in embryonic stem cells (ES), precipitated tumorigenesis after transplantation, i.e., 0% tumor-free outcomes at 3 × 105 cells/heart (light and dark green). For each condition, n indicates the number of treated animals. (B and C) TNF-α priming achieved tumor-free incorporation of transplanted stem cells (3 × 106 cells/heart; top panel), with embryonic stem cell–derived cardiomyocytes integrating with host myocardium as determined by CFP expression (B, bottom and C). Bars: (B, top) 2 mm; (B, bottom and C) 10 μm. (D) Transgenic TNF-α overexpression (TNFα-TG) up-regulated myocardial p38 MAPK and TGF-β expression with α-actinin used as a control for protein loading. (E and F) Blockade of the TGF-β superfamily through overexpression of Noggin or ΔTGFβRII precipitated tumor formation because of uncontrolled growth of implanted embryonic stem cells even at 3 × 105 cells/heart.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2118723&req=5

fig2: Neoplastic risk of embryonic stem cell therapy averted with transgenic cytokine overexpression. (A) Host heart (WT) demonstrated a finite capacity to secure tumor-free stem cell transplantation, i.e., 100% tumor-free outcome at 3 × 105 cells/heart versus 82% and 32% tumor-free outcomes at 106 cells/heart and 3 × 106 cells/heart, respectively (blue). Transgenic cardiac-restricted overexpression of TNF-α (TNF-TG) increased the capacity of host heart to guide stem cell cardiogenesis ifying uncontrolled growth, i.e., 100% tumor-free outcome at 3 × 106 cells/heart (red). Conversely, deletion of the kinase domain of the TGF-β receptor (ΔTGFβRII) or overexpression of the BMP inhibitor Noggin, both disrupting cardiogenic signaling in embryonic stem cells (ES), precipitated tumorigenesis after transplantation, i.e., 0% tumor-free outcomes at 3 × 105 cells/heart (light and dark green). For each condition, n indicates the number of treated animals. (B and C) TNF-α priming achieved tumor-free incorporation of transplanted stem cells (3 × 106 cells/heart; top panel), with embryonic stem cell–derived cardiomyocytes integrating with host myocardium as determined by CFP expression (B, bottom and C). Bars: (B, top) 2 mm; (B, bottom and C) 10 μm. (D) Transgenic TNF-α overexpression (TNFα-TG) up-regulated myocardial p38 MAPK and TGF-β expression with α-actinin used as a control for protein loading. (E and F) Blockade of the TGF-β superfamily through overexpression of Noggin or ΔTGFβRII precipitated tumor formation because of uncontrolled growth of implanted embryonic stem cells even at 3 × 105 cells/heart.
Mentions: The propensity for tumorigenic outcome correlated with embryonic stem cell load (Fig. 2 A). The neoplastic threat associated with stem cell transplantation was eliminated by cardiac-restricted transgenic expression of the stress cytokine TNF-α (TNF-α–TG), averting tumorigenic outcome even at doses of 10,000 cells per mg of myocardial tissue (∼3 × 106 stem cells/heart) that produced uncontrolled growth in wild-type hearts (Fig. 2 A). Up-regulated myocardial expression of TNF-α secured cardiac differentiation of implanted embryonic stem cells with proper integration within host myocardium occurring over an increasing range of stem cell loads (Fig. 2, A and B). Autofluorescence and cell fusion was ruled out by multiwavelength confocal visualization after immunohistochemical and nuclear probing (Fig. 2 C). The in vivo action of TNF-α was associated with myocardial up-regulation of the p38 mitogen-activated protein kinase and enhanced expression of the cardiogenic growth factor TGF-β (Fig. 2 D). Deletion of the TGF-β receptor kinase domain (ΔTGF-βRII) in embryonic stem cells or disruption of the stem cell ability to recognize members of the TGF-β superfamily, i.e., BMP through overexpression of the BMP inhibitor, noggin, prevented cardiac differentiation leading to tumor formation even at low stem cell loads (Fig. 2 A, E, and F). Thus, the tumorigenic risk of embryonic stem cells can be blunted by TNF-α cardiac-restricted overexpression or exaggerated by removal of the TGF-β superfamily guidance of transplanted cells.

Bottom Line: Here, the tumorigenic threat associated with embryonic stem cell transplantation was suppressed by cardiac-restricted transgenic expression of the reprogramming cytokine TNF-alpha, enhancing the cardiogenic competence of recipient heart.Characterized by a down-regulation of oncogenic markers, up-regulation, and nuclear translocation of cardiac transcription factors, this predetermined population yielded functional cardiomyocyte progeny.Thus, cardiopoietic programming establishes a strategy to hone stem cell pluripotency, offering a tumor-resistant approach for regeneration.

View Article: PubMed Central - PubMed

Affiliation: Marriott Heart Disease Research Program, Division of Cardiovascular Diseases, Department of Medicine, Mayo Clinic, Rochester, MN 55905, USA.

ABSTRACT
Embryonic stem cells have the distinct potential for tissue regeneration, including cardiac repair. Their propensity for multilineage differentiation carries, however, the liability of neoplastic growth, impeding therapeutic application. Here, the tumorigenic threat associated with embryonic stem cell transplantation was suppressed by cardiac-restricted transgenic expression of the reprogramming cytokine TNF-alpha, enhancing the cardiogenic competence of recipient heart. The in vivo aptitude of TNF-alpha to promote cardiac differentiation was recapitulated in embryoid bodies in vitro. The procardiogenic action required an intact endoderm and was mediated by secreted cardio-inductive signals. Resolved TNF-alpha-induced endoderm-derived factors, combined in a cocktail, secured guided differentiation of embryonic stem cells in monolayers produce cardiac progenitors termed cardiopoietic cells. Characterized by a down-regulation of oncogenic markers, up-regulation, and nuclear translocation of cardiac transcription factors, this predetermined population yielded functional cardiomyocyte progeny. Recruited cardiopoietic cells delivered in infarcted hearts generated cardiomyocytes that proliferated into scar tissue, integrating with host myocardium for tumor-free repair. Thus, cardiopoietic programming establishes a strategy to hone stem cell pluripotency, offering a tumor-resistant approach for regeneration.

Show MeSH
Related in: MedlinePlus