Key Event Title
Key Event Component
Key Event Overview
AOPs Including This Key Event
Level of Biological Organization
|Rodentia sp. 09ZR5#43||Rodentia sp. 09ZR5#43||Strong||NCBI|
How This Key Event Works
A large number of quantitative models have mathematically evaluated the increases in the number and volume of phenotypically altered (e.g. GSTP-positive) foci (Andersen et al., 1993, 1995; Andersen and Conolly, 1998; Bock, and Kohle, 2005; Conolly, and Andersen, 1997; Kim et al., 2003; Mills and Andersen, 1993; Moolgavkar et al., 1996; Portier et al., 1993, 1996). Thresholds for tumour promotion have been described for an increase in volume fraction of altered hepatic foci (Maronpot et al., 1993; Pitot et al., 1987; Teeguarden et al., 1999; Viluksela et al., 2000).
However, it should be noted that tumour promotion protocols create artificial circumstances by greatly accelerating the initiation rate of altered hepatic foci. In contrast, with sustained AHR activation, it is the development of altered hepatic foci (AHF) that naturally occur that are a primary target of the AHR-induced tumour promotion. AHF occur spontaneously in rats, albeit at a low rate. AHF may occur from either transformation of hepatocytes or of liver stem cells that have migrated to the liver parenchyma (Kuhlmann and Peschke, 2006). The rate of growth of AHF seems to be increased in TCDD-treated rats thereby demonstrating that sustained AHR activation either suppresses intrafocal apoptosis or increases intrafocal cell proliferation (Schrenk et al., 1994, 2004). The use of a nitrosamine may, or may not, create mutations in genes that dioxins would normally influence in a two-year cancer bioassay. In any case, the initiation-promotion data coupled to the understanding of the otherwise slow rate of emergence and growth of foci suggests that a critical number and volume of foci are necessary for TCDD-induced sustained AHR activation to promote rodent liver tumours. The dependency on the development of sufficient number and volume of altered hepatic foci may be a ModF for when the MIE actually occurs.
The timing of apoptosis inhibition within foci and increased cell proliferation has been discussed above. The results from initiation-promotion studies illustrate that TCDD-mediated reductions in intrafocal apoptosis can occur within months with continuous TCDD administration (Luebeck et al., 2000). Increased cell proliferation, which appears to occur more selectively in non-parenchymal cells, likewise ensues months after continuous TCDD exposure, suggesting that the observed periportal increase in labelling index occurs in hepatoblasts, biliary cells and other non-parenchymal cells such as oval cells (Hailey et al., 2005; Buchmann et al., 1994; Maronpot et al., 1993).
Hepatocellular adenomas may arise from clonal expansion within altered hepatic foci and cholangiolarcarcinomas from spontaneously initiated biliary progenitor cells near the canals of Hering. Prolonged administration of indole-3-carbinol, an AHR agonist found in broccoli, increased the number and volume of GST-P+ altered hepatic foci in a 26 week initiation-promotion study and hepatocellular adenomas were observed in some animals (Yamamoto et al., 2013). In this study, male F-344 rats received a diet containing 0.5% I3C and assuming they consumed 15% of their body weight daily and weighed on average 300 g, the daily dose of I3C is estimated at 750 mg/kg/day whereas in vivo gene expression data indicated a half-maximal dose of about 100 mg/kg for CYP1A2 mRNA (Coe et al., 2006).
At the organ level (next section), the occurrence of toxic hepatopathy and associated hepatocyte damage likely involves the development of a proliferative response by liver stem cells. Normal replacement of hepatocytes occurs by cell division of other, likely younger hepatocytes. However, in the case of extensive liver damage, replacement likely also occurs by stem cell proliferation (Tanaka et al., 2011). Wnt/β-catenin signalling plays a role in liver regeneration in many species (Nekak-Bowen and Monga, 2011) and may be stimulated by sustained AHR-activation. Sustained activation of the AHR by TCDD downregulated Wnt/β-catenin target genes in WB-F344 cells, considered a model for liver stem cells. Cell-to-cell interactions may also contribute to increased stem cell proliferation, The downregulation of β-catenin reduced cell adhesion and E-cadherin-mediated cell-cell junctions (Procházková et al., 2011a). Indirubin is an endogenous compound that can act as a weak AHR agonist. In WB-F344 cells, considered a model for liver stem/progenitor cells, doses of 100 pM indirubin increased gene expression of plakoglobin, a protein related to cadherin-mediated intracellular communication; however, doses of 1 or 10 μM indirubin decreased the expression of plakoglobin to a similar extent as did 5nM TCDD (Procházková et al., 2011b). For example, the regenerative properties of liver stem cells are modulated by stellate cells, which contain up to 80% of the vitamin A in the body (Senoo et al., 2010; Pintilie et al 2010).
The possibility that hepatic stem cells give rise to both tumor types cannot be discounted. Significant liver toxicity with damage to the parenchymal cells occurs prior to tumor formation and therefore, stem cell proliferation leading to development of both hepatic and biliary tumours seems likely. AHR-mediated transformation and associated changes in cell-cell contact and/or intercellular communication could enable stem cells to migrate from the progenitor cell niche to the liver parenchyma and there become an AHF through clonal expansion.
Several different methods have been established for obtain clonal populations of liver stem cells from rodents that are capable of differentiating into both hepatic and biliary cells (Sahin et al., 2008; Yavorkovsky et al., 1995; Fougere-Deschatrette et al., 2006). These cell lines could be used to develop assays for known AHR agonists to determine their potential effects on stem cell proliferation and biology.
How It Is Measured or Detected
Methods that have been previously reviewed and approved by a recognized authority should be included in the Overview section above. All other methods, including those well established in the published literature, should be described here. Consider the following criteria when describing each method: 1. Is the assay fit for purpose? 2. Is the assay directly or indirectly (i.e. a surrogate) related to a key event relevant to the final adverse effect in question? 3. Is the assay repeatable? 4. Is the assay reproducible?