Lentiviral reprogramming of human umbilical cord-derived mesenchymal stromal cells towards hepatocyte-like cells

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Scope of the method

The Method relates to
  • Human health
The Method is situated in
  • Basic Research
Type of method
  • In vitro - Ex vivo
This method makes use of
  • Human derived cells / tissues / organs
Specify the type of cells/tissues/organs
umbilical cord

Description

Method keywords
  • human adult stem cells
  • hepatocyte-like cells
  • lentiviral reprogramming
  • umbilical cord
Scientific area keywords
  • in vitro liver model
  • Drug-induced liver injury (DILI)
  • human hepatocyte-like cells
  • liver enriched transcription factor
  • human adult stem cells
Method description

Human umbilical cord-derived mesenchymal stromal cells (hUC-MSCs) express several key liver-specific transcription factors as well as hepatic progenitor markers. However, they still lack the hepatocyte nuclear factors 1-alpha (HNF1a) and 4-alpha (HNF4a), indispensable for their reprogramming towards hepatocyte-like cells. This method comprises the reprogramming of hUC-MSCs towards hepatocyte-like cells through HNF1a lentiviral over-expression. Whole genome microarray analysis revealed that the expression of the nuclear receptor retinoid X receptor (RXR) gamma and the nuclear transcription factor HNF4a, in HNF1a-transduced hUC-MSCs, was significantly upregulated compared to the control conditions. This expression was even higher than found in human hepatocytes. The same was observed for the uridine 5'-diphospho-glucuronosyltransferase (UGT) 1A family. Further, a significant upregulation was observed for alpha-foetoprotein (AFP), alpha1-antitrypsin (A1-AT), the phase I biotransformation enzymes cytochrome P450 (CYP) 1A2 and CYP2A6 and the drug transporter multidrug resistance protein (MDR) 1.

Lab equipment
  • Incubator (37 ± 1°C, 90 ± 5% humidity, 5.0 ± 1% CO2/air);
  • Type 2 laminar airflow HEK293T cells;
  • Ultracentrifuge Water bath (37 ± 1°C);
  • Inverse-phase contrast microscope;
  • Pipettes;
  • Pipettors;
  • Colorimetric reverse transcriptase assay;
  • Human hepatocytes;
  • Affymetrix microarray technologies;
  • Partek Genomics Suite Software.
Method status
  • Still in development

Pros, cons & Future potential

Advantages
  • Persistent expression of HNF1a transcription factor in hUC-MSCs;
  • Endogenous induction of HNF4a expression.
Challenges

Genomic integration of the lentiviral vector.

Modifications

Usage of a non-integrative reprogramming method e.g. mRNA transfection.

Future & Other applications

Generation of hepatocyte-like cells for the development of functional human liver-based in vitro models for pharmaco-toxicological purposes.

Contact person

Karolien Buyl

Organisations

Vrije Universiteit Brussel (VUB)
Pharmaceutical and Pharmacological Sciences
In Vitro Toxicology and Dermato-Cosmetology
Belgium
Brussels Region