Hepatocytes have a unique and unlimited ability for division allowing the regeneration of the liver after surgical resection or following chemical or viral insult. Taking the example of rat, it is known that 24 hours after a 70% partial hepatectomy, close to 90% of the hepatocytes in the remaining 30% are dividing while the full liver mass is expected to be restored within seven to ten days. Although the process is thought to be similar in humans, timing for DNA synthesis, cell division and full recovery may differ. Current evidence demonstrate a rapid regeneration during the first two weeks and a more gradual size increase over the next six-to-twelve months following a major hepatectomy. The very early events associated with the regenerative process are still being investigated and a complete sequence of proceedings is yet to be described. In rat, an increase in urokinase-like plasminogen activator (uPA) and uPA receptor (uPAR) is observed as early as one minute following hepatectomy and expression of this critical initiator of the metalloproteinase cascade keeps on increasing for at least one hour. Alteration of the extracellular matrix resulting from this surge in uPA is thought to be essential for the priming of the division of the hepatocytes and later on, of the other liver cell types. The Wnt/β-catenin pathway is also thought to be associated with early molecular events. A transient increase in β-catenin levels and a translocation to the nucleus take place five minutes after hepatectomy in rats. Return to normal β-catenin levels occurs two-to-three days later. The sudden over-expression of this transcription factor may help with the expression of not only uPAR but also other key transcription factors and cell cycle proteins such as c-myc and cyclin D1, respectively. Evidently, this series of biological changes is critical to the initiation of the regenerative process and the commitment of the hepatocytes to renew. However, very little is known about the exact source of these new hepatocytes and a more in-depth characterization of the molecular mechanisms is required for a complete understanding of the renewing ability of the liver.
Axin-2 is a known regulator of the stability of β-catenin in the Wnt signaling pathway and lineage tracing of Axin-2-positive cells has previously permitted the identification of stem cell niches in mammalian tissues.
Dr. Bruce Wang and colleagues from Stanford University School of Medicine used a similar lineage tracing approach of Axin-2-positive cells to identify the cell population responsible for the renewal of hepatocytes, localize its anatomical niche and decipher early molecular switches that control its activity. Thanks to a tamoxifen-inducible Axin-2-GFP mouse model, they identified a unique cell population of Axin-2-positive cells located around the central vein and were able to trace these cells over time. After one year, the marking of these Axin-2-positive cells remained and demonstrated their spreading towards the portal vein across the liver organ. Furthermore, all labeled cells expressed specifically the hepatocyte marker HNF4α but not markers of other liver cell types. Using the same technique of lineage tracing, they showed that these Axin-2-positive cells located near the central vein were able to self-renew arising from pre-existing Axin-2-positive cells. Tbx3, a key transcription factor known to maintain a state of pluripotency in stem cells, was shown by RNAseq to be significantly up-regulated in Axin-2-positive cells when compared to Axin-2-negative cells. Similarly to stem cells and contrary to most mature hepatocytes, Axin-2-positive cells are diploid, which is a property necessary for indefinite duplication. Finally, expression of Axin-2 near the central vein suggests a source of Wnt in the microenvironment of these cells. Normal liver was screened for all nineteen Wnt proteins by RNA
in situ hybridization and results were validated by RT-qPCR. The authors observed the exclusive over-expression of both Wnt2 and Wnt9b in endothelial cells around the central vein, which may act along Axin-2-positive cells as a stem cell niche for future hepatocytes. By conditionally deleting Wntless, a Wnt-specific transporter required for Wnt secretion, they concluded that the secretion of these two Wnt proteins by endothelial cells at the central vein is essential for the preservation of the high-proliferative state of Axin-2-positive cells. Altogether, these results shed some light on the regenerative capacity of the liver and go a long way towards a better understanding of liver pathogenesis and most specifically, liver tumorigenesis.
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