Generation of Cardiomyocytes in Pipe-Based Microbioreactor Under Segmented Flow

Cell Physiol Biochem. 2016;38(5):1883-96. doi: 10.1159/000445550. Epub 2016 May 9.

Abstract

Background/aims: Embryonic stem (ES) cells have got a broad range differentiation potential. The differentiation is initiated via aggregation of non-differentiated ES cells into embryoid body (EB) capable of multi-lineage development. However experimental variables present in standard differentiation techniques lead to high EB heterogeneity, affecting development into the cells of desired lineage, and do not support the process automatization and scalability.

Methods: Here we present a novel pipe based microbioreactor (PBM) setup based on segmented flow, designed for spatial maintenance of temperature, nutrition supply, gas supply and sterility.

Results: We verified PBM feasibility for continuous process generating cardiac cells starting from single ES cell suspension followed by EB formation for up to 10 days. The ES cells used in the study were genetically modified for cardiac-specific EGFP expression allowing optical monitoring of cardiomyocytes while EBs remained within PBM for up to 10 days. Efficiency of cardiac cells formation within PBM was similar compared to a standard hanging drop based protocol.

Conclusion: Our findings ensure further development of microfluidic bioreactor technology to enable robust cardiomyocytes production for needs of drug screening, tissue engineering and other applications.

MeSH terms

  • Animals
  • Bioreactors
  • Cell Culture Techniques / instrumentation
  • Cell Culture Techniques / methods*
  • Cell Differentiation
  • Cell Line
  • Flow Cytometry
  • GATA4 Transcription Factor / genetics
  • GATA4 Transcription Factor / metabolism
  • Mice
  • Microfluidics / instrumentation
  • Microfluidics / methods
  • Microscopy
  • Mouse Embryonic Stem Cells / cytology*
  • Mouse Embryonic Stem Cells / metabolism
  • Myocytes, Cardiac / cytology*
  • Myocytes, Cardiac / metabolism
  • Nanog Homeobox Protein / genetics
  • Nanog Homeobox Protein / metabolism
  • RNA, Messenger / isolation & purification
  • Real-Time Polymerase Chain Reaction
  • Tissue Engineering

Substances

  • GATA4 Transcription Factor
  • Gata4 protein, mouse
  • Nanog Homeobox Protein
  • Nanog protein, mouse
  • RNA, Messenger