title: Enhanced gap junction expression in myoblast-containing engineered tissue.
abstract: Transplantation of skeletal myoblasts (SMs) has been investigated as a potential cardiac cell therapy approach. SM are available autologously, predetermined for muscular differentiation and resistant to ischemia. Major hurdles for their clinical application are limitations in purity and yield during cell isolation as well as the absence of gap junction expression after differentiation into myotubes. Furthermore, transplanted SMs do not functionally or electrically integrate with the host myocardium. Here, we describe an efficient method for isolating homogeneous SM populations from neonatal mice and demonstrate persistent gap junction expression in an engineered tissue. This method resulted in a yield of 1.4 × 10(8) high-purity SMs (>99% desmin positive) after 10 days in culture from 162.12 ± 11.85 mg muscle tissue. Serum starvation conditions efficiently induced differentiation into spontaneously contracting myotubes that coincided with loss of gap junction expression. For mechanical conditioning, cells were integrated into engineered tissue constructs. SMs within tissue constructs exhibited long term survival, ordered alignment, and a preserved ability to differentiate into contractile myotubes. When the tissue constructs were subjected to passive longitudinal tensile stress, the expression of gap junction and cell adherence proteins was maintained or increased throughout differentiation. Our studies demonstrate that mechanical loading of SMs may provide for improved electromechanical integration within the myocardium, which could lead to more therapeutic opportunities.
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Sureshkumar Perumal
authors.initials: SP
authors.lastname: Srinivasan
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Neef
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Treskes
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.
authors.forename: Oliver J
authors.initials: OJ
authors.lastname: Liakopoulos
authors.affiliation: Department of Cardiac and Thoracic and Vascular Surgery, German Heart Institute Berlin, Berlin, Germany.|Berlin-Brandenburg Center for Regenerative Therapies, Berlin, Germany.
authors.forename: Christof
authors.initials: C
authors.lastname: Stamm
authors.affiliation: Department of Anesthesiology, Perioperative and Pain Medicine, Children's Hospital Boston and Harvard Medical School, Boston, MA, USA.
authors.forename: Douglas B
authors.initials: DB
authors.lastname: Cowan
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.
authors.forename: Navid
authors.initials: N
authors.lastname: Madershahian
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.
authors.forename: Elmar
authors.initials: E
authors.lastname: Kuhn
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.
authors.forename: Ingo
authors.initials: I
authors.lastname: Slottosch
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wittwer
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wahlers
authors.affiliation: Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.|Center for Molecular Medicine, University of Cologne, Cologne, Germany.
authors.forename: Yeong-Hoon
authors.initials: YH
authors.lastname: Choi
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D002469:Cell Separation / Q000379:methods*; D017629:Gap Junctions / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D032448:Myoblasts, Skeletal / Q000166:cytology*; D023822:Tissue Engineering; D016474:Weight-Bearing