Hepatic differentiation of human embryonic stem cells on microcarriers

J Biotechnol. 2014 Mar 20;174:39-48. doi: 10.1016/j.jbiotec.2014.01.025. Epub 2014 Jan 27.

Abstract

Translation of stem cell research to industrial and clinical settings mostly requires large quantities of cells, especially those involving large organs such as the liver. A scalable reactor system is desirable to ensure a reliable supply of sufficient quantities of differentiated cells. To increase the culture efficiency in bioreactor system, high surface to volume ratio needs to be achieved. We employed a microcarrier culture system for the expansion of undifferentiated human embryonic stem cells (hESCs) as well as for directed differentiation of these cells to hepatocyte-like cells. Cells in single cell suspension were attached to the bead surface in even distribution and were expanded to 1×10(6)cells/ml within 2 days of hESC culture with maintenance of the level of pluripotency markers. Directed differentiation into hepatocyte-like cells on microcarriers, both in static culture and stirred bioreactors, induced similar levels of hepatocyte-like cell differentiation as observed with cells cultured in conventional tissue culture plates. The cells expressed both immature and mature hepatocyte-lineage genes and proteins such as asialoglycoprotein receptor-1 (ASGPR-1) and albumin. Differentiated cells exhibited functional characteristics such as secretion of albumin and urea, and CYP3A4 activity could be detected. Microcarriers thus offer the potential for large-scale expansion and differentiation of hESCs induced hepatocyte-like cells in a more controllable bioreactor environment.

Keywords: Bioreactor; Hepatic differentiation; Human embryonic stem cells; Microcarrier; Suspension culture system.

MeSH terms

  • Albumins / genetics
  • Albumins / metabolism
  • Asialoglycoprotein Receptor / genetics
  • Asialoglycoprotein Receptor / metabolism
  • Biomarkers / metabolism*
  • Biomarkers / urine*
  • Bioreactors
  • Cell Culture Techniques / methods*
  • Cell Differentiation / physiology
  • Cell Growth Processes
  • Cell Line
  • Cytochrome P-450 CYP3A / genetics
  • Cytochrome P-450 CYP3A / metabolism
  • Embryonic Stem Cells / cytology*
  • Embryonic Stem Cells / metabolism
  • Hepatocytes / cytology*
  • Hepatocytes / metabolism
  • Humans

Substances

  • ASGR1 protein, human
  • Albumins
  • Asialoglycoprotein Receptor
  • Biomarkers
  • Cytochrome P-450 CYP3A
  • CYP3A4 protein, human