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Cell differentiation mediated by co-culture of human umbilical cord blood stem cells with murine hepatic cells

  • Maria StecklumEmail author
  • Annika Wulf-Goldenberg
  • Bettina Purfürst
  • Antje Siegert
  • Marlen Keil
  • Klaus Eckert
  • Iduna Fichtner
Article

Abstract

In the present study, purified human cord blood stem cells were co-cultivated with murine hepatic alpha mouse liver 12 (AML12) cells to compare the effect on endodermal stem cell differentiation by either direct cell-cell interaction or by soluble factors in conditioned hepatic cell medium. With that approach, we want to mimic in vitro the situation of preclinical transplantation experiments using human cells in mice. Cord blood stem cells, cultivated with hepatic conditioned medium, showed a low endodermal differentiation but an increased connexin 32 (Cx32) and Cx43, and cytokeratin 8 (CK8) and CK19 expression was monitored by reverse transcription polymerase chain reaction (RT-PCR). Microarray profiling indicated that in cultivated cord blood cells, 604 genes were upregulated 2-fold, with the highest expression for epithelial CK19 and epithelial cadherin (E-cadherin). On ultrastructural level, there were no major changes in the cellular morphology, except a higher presence of phago(ly)some-like structures observed. Direct co-culture of AML12 cells with cord blood cells led to less incisive differentiation with increased sex-determining region Y-box 17 (SOX17), Cx32 and Cx43, as well as epithelial CK8 and CK19 expressions. On ultrastructural level, tight cell contacts along the plasma membranes were revealed. FACS analysis in co-cultivated cells quantified dye exchange on low level, as also proved by time relapse video-imaging of labelled cells. Modulators of gap junction formation influenced dye transfer between the co-cultured cells, whereby retinoic acid increased and 3-heptanol reduced the dye transfer. The study indicated that the cell-co-cultured model of human umbilical cord blood cells and murine AML12 cells may be a suitable approach to study some aspects of endodermal/hepatic cell differentiation induction.

Keywords

Co-culture Murine hepatic cells (AML12) Umbilical cord blood cells (UCBC) Cell differentiation 

Notes

Acknowledgments

The skilful technical assistance of Mrs. Vannauer is gratefully acknowledged. We thank the HELIOS-Clinics GmbH in Berlin-Buch and the Vivantes Clinics in Berlin for collecting and providing the umbilical cord blood. The work was supported by a grant from the Federal Ministry of Education and Research (BMBF; 01GN0528)

Supplementary material

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11626_2014_9817_MOESM2_ESM.doc (54 kb)
ESM 2 (DOC 54 kb)

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Copyright information

© The Society for In Vitro Biology 2014

Authors and Affiliations

  • Maria Stecklum
    • 1
    • 3
    Email author
  • Annika Wulf-Goldenberg
    • 3
  • Bettina Purfürst
    • 1
  • Antje Siegert
    • 3
  • Marlen Keil
    • 1
    • 2
  • Klaus Eckert
    • 1
    • 3
  • Iduna Fichtner
    • 1
  1. 1.Max Delbrück Center for Molecular Medicine, Berlin-BuchBerlinGermany
  2. 2.Charite—Universitätsmedizin Berlin, Campus Virchow KlinikumBerlinGermany
  3. 3.EPO GmbHBerlinGermany

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