Liver regeneration through progenitor cell response : role of the microenvironment in a murine model of chronic liver injury

Rapidly after liver damage, unharmed hepatocytes divide en masse to compensate for the endured cell loss and regain normal function. However, in case of massive and/or chronic injury this process is insufficient due to either paucity of hepatocytes able to engage into the regenerative process or replicative inability of the remaining hepatocytes. In those conditions, a dormant compartment of progenitor cells is activated and considered as a rescue mechanism for functional liver mass regeneration. Liver progenitor cells (LPC) are described as bipotential being able to differentiate into biliary or hepatocytic lineages, depending on the injurious process. The accompanying microenvironment, consisting of hepatic stellate cells (HSC) /myofibroblasts (MF), Kupffer cells (KC), extracellular matrix (ECM) and soluble factors is believed to have a major role in the regulation and modulation of the LPC response. The aim of the present PhD thesis was to evaluate the contribution of this microenvironment to the liver progenitor cell response induced in mice after administration of a choline deficient diet supplemented in ethionine (CDE diet). In a first study using the CDE-model, we designed a time line set up whereby mice were sacrificed at certain time points in order to determine the chronology of the events taking place during LPC-mediated liver regeneration. Hereby, we were able to demonstrate that HSC/MF are rapidly activated (within three days) after the initiation of the diet. These mesenchymal cells were found to proliferate and to deposit ECM around the portal area before the appearance of LPC. Several days later, LPC started to proliferate and to form branching structures while elongating from the pre-existing canals of Hering, the connective structures between hepatocytic canaliculi and bile ductules, and invading the parenchyma. During this parenchymal invasion, we documented that the HSC/MF were wrapped around the LPC. Moreover, the HSC/MF and their ECM deposition were preceding the LPC at the edges of migration. In addition, the quantity of HSC/MF/ECM was shown to increase in parallel with the accumulation of LPC. Taken together, the mesenchymal contribution (both HSC/MF and ECM) seems to be a requisite for the appearance, the proliferation and the migration of the LPC during this attempt of regeneration. In a following study, we focused on the role of resident liver macrophages or Kupffer cells (KC) in the LPC-mediated liver regeneration as inflammation is also accompanying the progenitor cell response. In normal liver, KC were uniformly scattered in the parenchyma. At day 3 of the CDE model, KC were preferentially located around the portal area. Later, KC were migrating from portal to central area (day 7) to be found mainly in the region around the central veins (day 10). This showed that activated KC are also preceding migrating LPC, seemingly even in advance of the HSC/MF. We postulated that KC could potentially influence the LPC response and investigated this by depleting the macrophage population in the CDE model. Removal of the KC did not influence the proliferative capacity of the LPC. Instead, this had an effect on the migratory phenotype of the LPC, which were found to be rounder, strongly interconnected and closer to the portal tract in comparison to non-depleted controls. In addition, some LPC of KC-depleted animals appeared organised in pseudo-ducts leading to the suggestion that these cells were in a more differentiated stage and leaning towards a biliary phenotype. This was in contrast to KC-containing CDE progenitor cells which were arranged in filaments without notable differentiation. KC are therefore documented to play a role not in the activation or proliferation of the LPC but in their migration and differentiation.(SBIM 3) -- UCL, 201