Hypoxic preconditioning potentiates the trophic effects of mesenchymal stem cells on co-cultured human primary hepatocytes

Stem Cell Res Ther. 2015 Dec 1;6:237. doi: 10.1186/s13287-015-0218-7.


Introduction: Mesenchymal stem/stromal cells (MSCs) improve the metabolic function of co-cultured hepatocytes. The present study aimed to further enhance the trophic effects of co-culture with hepatocytes using hypoxic preconditioning (HPc) of the MSCs and also to investigate the underlying molecular mechanisms involved.

Methods: Human adipose tissue-derived MSCs were subjected to hypoxia (2 % O2; HPc) or normoxia (20 % O2) for 24 h and then co-cultured with isolated human hepatocytes. Assays of metabolic function and apoptosis were performed to investigate the hepatotrophic and anti-apoptotic effects of co-culture. Indirect co-cultures and co-culture with MSC-conditioned medium investigated the role of paracrine factors in the hepatotrophic effects of co-culture. Reactive oxygen species (ROS) activity was antagonised with N-acetylcysteine to investigate whether HPc potentiated the effects of MSCs by intracellular ROS-dependent mechanisms. Tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and extracellular collagen production was determined and CASP9 and BAX/BCL-2 signalling pathways analysed to investigate the role of soluble factors, extracellular matrix deposition, and apoptosis-associated gene signalling in the effects of co-culture.

Results: HPc potentiated the hepatotrophic and anti-apoptotic effects of co-culture by ROS-dependent mechanisms. There was increased MSC TGF-β1 production, and enhanced MSC deposition of extracellular collagen, with reduced synthesis of TNF-α, as well as a downregulation of the expression of pro-apoptotic CASP9, BAX, BID and BLK genes and upregulated expression of anti-apoptotic BCL-2 in hepatocytes.

Conclusions: HPc potentiated the trophic and anti-apoptotic effects of MSCs on hepatocytes via mechanisms including intracellular ROS, autocrine TGF-β, extracellular collagen and caspase and BAX/BCL-2 signalling pathways.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / genetics
  • Cell Hypoxia
  • Coculture Techniques
  • Collagen / metabolism
  • Culture Media, Conditioned
  • Down-Regulation
  • Genes, bcl-2
  • Hepatocytes / cytology*
  • Hepatocytes / metabolism*
  • Hepatocytes / transplantation
  • Humans
  • Ischemic Preconditioning
  • Mesenchymal Stem Cell Transplantation
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / metabolism*
  • Paracrine Communication
  • Reactive Oxygen Species / metabolism
  • Transforming Growth Factor beta1 / metabolism
  • Tumor Necrosis Factor-alpha / metabolism
  • bcl-2-Associated X Protein / genetics


  • BAX protein, human
  • Culture Media, Conditioned
  • Reactive Oxygen Species
  • Transforming Growth Factor beta1
  • Tumor Necrosis Factor-alpha
  • bcl-2-Associated X Protein
  • Collagen