Enrichment of cardiogenic cell populations from murine skeletal muscle

Yeong-Hoon Choi, Klaus Neef, Philipp Treskes, Sureshkumar Perumal Srinivasan, Christof Stamm, Pb Rahmanian, Oliver J Liakopoulos, Thorsten Wittwer, Thorsten Wahlers

Research output: Contribution to conferenceAbstractpeer-review

Abstract

Introduction: Adult stem cells with regenerative potential for cardiac disease, derived from the patient, represent an ideal source for cell therapeutic approaches aiming at replacing dysfunctional heart tissue after injury or disease. Here, we present data for the isolation of cell populations with cardiogenic potential from murine skeletal muscle.

Methods: A subpopulation of non-adherent cells was isolated from murine skeletal muscle by preplating and applying cell culture conditions favoring cluster formation. Cells were phenotypically analyzed by immunocytochemistry (IHC) and quantitative PCR (qPCR). Transplantations of 5×105 fluorescently labeled cells to a murine infarct model were performed and analyzed after one week immunohistochemically to locate the transplanted cells and determine integration into the host tissue and functional maturation.

Results: After mechanical and enzymatic dissociation of muscle tissue from ten neonatal mice and applying three successive pre-plating procedures 11.0±5.1×106 spontaneously contracting, cluster forming cells (ISH0) could be generated. Cluster sizes increased after further cell culture including hanging-drop treatment (H: 114.7±32.3µm) or incubation on a shaker platform (S: 121.2±22.5µm) as compared to static incubation (I: 68.2±21.5µm; p<0.01 vs. H and S). IHC analyses showed significant increase of cardiac markers α-actinin (I: 72.2±1.6%; S: 67.7±7.2%; H: 80.8±6.3%; all p<0.001 vs. ISH0: 32.6±6.2%) and troponin T (I: 55.8±3.9%; S: 67.4±10.3%; H: 82.7±5.3%; all p<0.001 vs. ISH0: 28.2±3.5%). This was confirmed by qPCR, which further revealed an increased expression of cardiac myosin heavy chain 6 (H: 40 fold, S: 17 fold, I: 15 fold increase vs. ISH0; all p<0.01). Histological analysis 1 week after cell transplantation revealed fluorescently labeled cells in the peri-infarct region. IHC staining for α-actinin showed typical striated structures of cardiomyocytes for transplanted cells. Staining for connexin 43 showed positive signals on the borders of transplanted cells.

Conclusions: Subpopulations of cells with cardiogenic potential can be isolated from skeletal muscle. Forcing those cells into clusters results in a pronounced shift of phenotypic properties from skeletal muscle towards cardiomyocytes. This cell population represents a highly promising source of autologously available, non-transgenic cells with potential for clinical application in regenerative therapies of the heart.
Original languageEnglish
Publication statusPublished - 12 Feb 2012
Event41st Annual Meeting of the German Society for Cardiovascular and Thoracic Surgery, - Freiburg, Germany
Duration: 12 Feb 201215 Feb 2012

Conference

Conference41st Annual Meeting of the German Society for Cardiovascular and Thoracic Surgery,
CountryGermany
CityFreiburg
Period12/02/1215/02/12

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