title: A Simplified, Langendorff-Free Method for Concomitant Isolation of Viable Cardiac Myocytes and Nonmyocytes From the Adult Mouse Heart.
abstract: BACKGROUND
  Cardiovascular disease represents a global pandemic. The advent of and recent advances in mouse genomics, epigenomics, and transgenics offer ever-greater potential for powerful avenues of research. However, progress is often constrained by unique complexities associated with the isolation of viable myocytes from the adult mouse heart. Current protocols rely on retrograde aortic perfusion using specialized Langendorff apparatus, which poses considerable logistical and technical barriers to researchers and demands extensive training investment.
  
  
  OBJECTIVE
  To identify and optimize a convenient, alternative approach, allowing the robust isolation and culture of adult mouse cardiac myocytes using only common surgical and laboratory equipment.
  
  
  RESULTS
  Cardiac myocytes were isolated with yields comparable to those in published Langendorff-based methods, using direct needle perfusion of the LV ex vivo and without requirement for heparin injection. Isolated myocytes can be cultured antibiotic free, with retained organized contractile and mitochondrial morphology, transcriptional signatures, calcium handling, responses to hypoxia, neurohormonal stimulation, and electric pacing, and are amenable to patch clamp and adenoviral gene transfer techniques. Furthermore, the methodology permits concurrent isolation, separation, and coculture of myocyte and nonmyocyte cardiac populations.
  
  
  CONCLUSIONS
  We present a novel, simplified method, demonstrating concomitant isolation of viable cardiac myocytes and nonmyocytes from the same adult mouse heart. We anticipate that this new approach will expand and accelerate innovative research in the field of cardiac biology.
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.).
authors: Matthew
authors: M
authors: Ackers-Johnson
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.).
authors: Peter Yiqing
authors: PY
authors: Li
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.).
authors: Andrew P
authors: AP
authors: Holmes
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.).
authors: Sian-Marie
authors: SM
authors: O'Brien
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.).
authors: Davor
authors: D
authors: Pavlovic
authors: From the Cardiovascular Research Institute, Centre for Translational Medicine MD6, National University Health System, Singapore (M.A.-J., P.Y.L., R.S.F.); Genome Institute of Singapore (M.A.-J., R.S.F.); and Institute of Cardiovascular Sciences, University of Birmingham, UK (A.P.H., S.-M.O., D.P.). mdcrfsy@nus.edu.sg.
authors: Roger S
authors: RS
authors: Foo
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002469:Cell Separation / Q000379:methods*; D018920:Coculture Techniques / Q000379:methods; D005347:Fibroblasts / Q000502:physiology*; D006352:Heart Ventricles / Q000166:cytology; D000067268:Isolated Heart Preparation / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D009206:Myocardium / Q000166:cytology*; D032383:Myocytes, Cardiac / Q000502:physiology*
keywords: Langendorff-free; cardiac fibroblasts; cardiomyocytes; cardiovascular disease; coculture; mouse models; single-cell isolation