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Dynamic Support Culture of Murine Skeletal Muscle-Derived Stem Cells Improves Their Cardiogenic Potential In Vitro.

Neef K, Treskes P, Xu G, Drey F, Srinivasan SP, Saric T, Nembo E, Semmler J, Nguemo F, Stamm C, Cowan DB, Deppe AC, Scherner M, Wittwer T, Hescheler J, Wahlers T, Choi YH - Stem Cells Int (2015)

Bottom Line: A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation.Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli.Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, Heart Center, University of Cologne, 50937 Cologne, Germany ; Center for Molecular Medicine Cologne, University of Cologne, 50931 Cologne, Germany.

ABSTRACT
Ischemic heart disease is the main cause of death in western countries and its burden is increasing worldwide. It typically involves irreversible degeneration and loss of myocardial tissue leading to poor prognosis and fatal outcome. Autologous cells with the potential to regenerate damaged heart tissue would be an ideal source for cell therapeutic approaches. Here, we compared different methods of conditional culture for increasing the yield and cardiogenic potential of murine skeletal muscle-derived stem cells. A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation. In contrast to static culture conditions, dynamic culture with or without previous hanging drop preculture led to significantly increased cluster diameters and the expression of cardiac specific markers on the protein and mRNA level. Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli. This data indicates that skeletal muscle-derived stem cells are capable of adopting enhanced cardiac muscle cell-like properties by applying specific culture conditions. Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells.

No MeSH data available.


Related in: MedlinePlus

Characteristics of cell clusters derived from MDSCs. (a) Representative phase contrast microphotographs of cell clusters from different cell culture conditions. While single contracting cells and smaller clusters were visible in ISH0 cultures, most cells were organized in clusters of loosely attached cells applying static (I) and dynamic (S and H) culture conditions at day 12 of cultivation (I12, S12, and H12). Scale: 100 μm. (b) Average sizes of clusters in initial ISH0 cell populations and after twelve days applying different culture conditions. ∗∗∗p < 0.001. (c) Total numbers of nonadherent cells obtained from muscles of 10 neonatal mice over the course of 12 days applying different culture conditions (n = 12). (d) Ratios of nonadherent cells over the course of 12 days applying different culture conditions (n = 12).
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fig2: Characteristics of cell clusters derived from MDSCs. (a) Representative phase contrast microphotographs of cell clusters from different cell culture conditions. While single contracting cells and smaller clusters were visible in ISH0 cultures, most cells were organized in clusters of loosely attached cells applying static (I) and dynamic (S and H) culture conditions at day 12 of cultivation (I12, S12, and H12). Scale: 100 μm. (b) Average sizes of clusters in initial ISH0 cell populations and after twelve days applying different culture conditions. ∗∗∗p < 0.001. (c) Total numbers of nonadherent cells obtained from muscles of 10 neonatal mice over the course of 12 days applying different culture conditions (n = 12). (d) Ratios of nonadherent cells over the course of 12 days applying different culture conditions (n = 12).

Mentions: Sizes of cell clusters (Figures 2(a) and 2(b)) after 12 days of static culture conditions did not differ significantly from initial ISH0 cell clusters (ISH0: 66.4 ± 2.0 μm, n = 9; I12: 68.2 ± 5.2 μm, n = 5), but clusters were significantly larger under both dynamic conditions (S12: 121.2 ± 3.9 μm; H12: 114.7 ± 5.2 μm; both p < 0.001 versus ISH0 and I12). Development of the number of nonadherent cells counted in suspension over the course of 12 days did not differ significantly among I, S, and H and showed a loss of approximately 6% of nonadherent cells per day, resulting in 28.4% (I: 3.13 ± 2.36 × 106 cells), 19.4% (S: 2.14 ± 1.34 × 106 cells), and 17.3% (H: 1.9 ± 1.53 × 106 cells) of the original ISH0 cells remaining after 12 days (Figure 2(c)). Although cell numbers continually declined during 12 days of static or dynamic culture (Figure 2(c)) an increase in ratios of nonadherent cells was observed (Figure 2(d)).


Dynamic Support Culture of Murine Skeletal Muscle-Derived Stem Cells Improves Their Cardiogenic Potential In Vitro.

Neef K, Treskes P, Xu G, Drey F, Srinivasan SP, Saric T, Nembo E, Semmler J, Nguemo F, Stamm C, Cowan DB, Deppe AC, Scherner M, Wittwer T, Hescheler J, Wahlers T, Choi YH - Stem Cells Int (2015)

Characteristics of cell clusters derived from MDSCs. (a) Representative phase contrast microphotographs of cell clusters from different cell culture conditions. While single contracting cells and smaller clusters were visible in ISH0 cultures, most cells were organized in clusters of loosely attached cells applying static (I) and dynamic (S and H) culture conditions at day 12 of cultivation (I12, S12, and H12). Scale: 100 μm. (b) Average sizes of clusters in initial ISH0 cell populations and after twelve days applying different culture conditions. ∗∗∗p < 0.001. (c) Total numbers of nonadherent cells obtained from muscles of 10 neonatal mice over the course of 12 days applying different culture conditions (n = 12). (d) Ratios of nonadherent cells over the course of 12 days applying different culture conditions (n = 12).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig2: Characteristics of cell clusters derived from MDSCs. (a) Representative phase contrast microphotographs of cell clusters from different cell culture conditions. While single contracting cells and smaller clusters were visible in ISH0 cultures, most cells were organized in clusters of loosely attached cells applying static (I) and dynamic (S and H) culture conditions at day 12 of cultivation (I12, S12, and H12). Scale: 100 μm. (b) Average sizes of clusters in initial ISH0 cell populations and after twelve days applying different culture conditions. ∗∗∗p < 0.001. (c) Total numbers of nonadherent cells obtained from muscles of 10 neonatal mice over the course of 12 days applying different culture conditions (n = 12). (d) Ratios of nonadherent cells over the course of 12 days applying different culture conditions (n = 12).
Mentions: Sizes of cell clusters (Figures 2(a) and 2(b)) after 12 days of static culture conditions did not differ significantly from initial ISH0 cell clusters (ISH0: 66.4 ± 2.0 μm, n = 9; I12: 68.2 ± 5.2 μm, n = 5), but clusters were significantly larger under both dynamic conditions (S12: 121.2 ± 3.9 μm; H12: 114.7 ± 5.2 μm; both p < 0.001 versus ISH0 and I12). Development of the number of nonadherent cells counted in suspension over the course of 12 days did not differ significantly among I, S, and H and showed a loss of approximately 6% of nonadherent cells per day, resulting in 28.4% (I: 3.13 ± 2.36 × 106 cells), 19.4% (S: 2.14 ± 1.34 × 106 cells), and 17.3% (H: 1.9 ± 1.53 × 106 cells) of the original ISH0 cells remaining after 12 days (Figure 2(c)). Although cell numbers continually declined during 12 days of static or dynamic culture (Figure 2(c)) an increase in ratios of nonadherent cells was observed (Figure 2(d)).

Bottom Line: A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation.Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli.Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, Heart Center, University of Cologne, 50937 Cologne, Germany ; Center for Molecular Medicine Cologne, University of Cologne, 50931 Cologne, Germany.

ABSTRACT
Ischemic heart disease is the main cause of death in western countries and its burden is increasing worldwide. It typically involves irreversible degeneration and loss of myocardial tissue leading to poor prognosis and fatal outcome. Autologous cells with the potential to regenerate damaged heart tissue would be an ideal source for cell therapeutic approaches. Here, we compared different methods of conditional culture for increasing the yield and cardiogenic potential of murine skeletal muscle-derived stem cells. A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation. In contrast to static culture conditions, dynamic culture with or without previous hanging drop preculture led to significantly increased cluster diameters and the expression of cardiac specific markers on the protein and mRNA level. Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli. This data indicates that skeletal muscle-derived stem cells are capable of adopting enhanced cardiac muscle cell-like properties by applying specific culture conditions. Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells.

No MeSH data available.


Related in: MedlinePlus