In the liver, hepatic stem/progenitor cells (HPCs) represent a facultative stem/progenitor cell compartment located within canals of Hering and bile ductules. HPCs are bipotent stem cells characterized by the capability to differentiate towards mature hepatocytes and cholangiocytes. In human Non Alcoholic Fatty Liver Disease (NAFLD), HPCs can activate and proliferate as a consequence of impaired proliferation of mature hepatocyes due to lipotoxicity. Recently, a niche of stem cells endowed within peribiliary glands (PBGs) has been described along extrahepatic and large intrahepatic bile ducts. This heterogeneous population of stem and progenitor cells has been collectively named Biliary Tree Stem Cells (BTSCs) and is implicated in the pathogenesis of human cholangiopathies.The general aim of the present project will be to define the role and the activation mechanisms of HPCs and BTSCs in human liver diseases. Specific objectives of the present project will be: i) to investigate the activation of HPCs in the progression of NAFLD and the relationship with levels of Lipopolysaccharide (LPS), platelets and macrophage activation; ii) to study the signaling pathways driving the BTSC activation in human Primary Sclerosing Cholangitis (PSC) and Non anastomotic strictures (NAS) in transplanted livers; ii) to individuate the role of peribiliary glands (PBGs) in biliary regeneration and in the pathogenesis of biliary damage by a lineage tracing approach in mice.
Data obtained from this project will lead i) to understand cellular mechanisms at the basis of NAFLD progression; ii) to characterize the anatomy and physiology of PBG stem cell niche, its behavior in regenerative response, and the signals involved in niche activation; iii) to clarify the pathogenesis of cholangiopathies and biliary fibrosis; iv) to individuate novel molecular tools and therapeutic approaches targeting stem cell niches.
The present project will clarify important aspects on the role of HPCs andBTSCs within the liver and the relevant pathways implicated in the activation of stem/progenitor cell niches. This project will examine regenerative pathways in several human pathologies and will examine their role in the response to pathologic conditions. Moreover, this project will individuate the role of duodenal cells as a candidate source for regenerative medicine of the liver.
The objective #1 will characterize mechanisms which can influence the development of NASH, fibrosis and cirrhosis. Moreover, it will clarify the contribution of TLRs pathways in macrophage activation and its role in the evolution to NASH; Will examine whether TLR4 is triggered by Escherichia Coli-derived LPS levels arriving to the liver via the portal system as the result of bacterial translocation from the gut. Many experimental and clinical evidence suggest that gut microbiota may be implicated in NAFLD. Experimental studies drew the attention on the role of LPS from gut microbiota in favoring the occurrence of NASH. The interplay between LPS and human NAFLD is less clear. Therefore, our study would test whether NAFLD is associated with enhanced LPS uptake by hepatic cells with potential effects in terms of inflammation and eventually hepatic fibrosis. We will evaluate the platelets role and their activation on NASH and fibrosis. Finally, our study will elucidate whether treatment with aspirin or modulation of TLR4 pathway would be able to reduce the progression to fibrosis. This part will have important implications in the clinics by further characterizing the liver-gut axis and its perturbation in NAFLD patients with the possibility to individuate novel therapeutic targets. The correlation of histological aspects with serum levels of LPS will furnish relevant insight, and data obtained from this part of the project will be useful to understand cellular mechanisms at the basis of NAFLD progression and could individuate novel molecular tools and cellular targets for therapeutic approaches.
The objective #2 will further clarify the pathways involved in biliary regeneration during cholangiopathies. In particular, the activation of HPC niche due to cholangiocyte loss in PBC patients will be examined in human and in rodent experimental models. HPC activation and DR will be put in the context of a deep clinical characterization of our PBC patient cohort to individuate novel histological features useful for risk stratification and to predict response to first-line therapy (i.e. UDCA). This study will look at the mechanisms leading to extension of DR with particular regard to the loss of ductular-canalicular junction. This objective of the project will further characterize pathogenetic mechanisms underlying biliary complications in transplanted organs and assess whether they are related to ischemia. In this setting, the role of micro-vessels and VEGF system will be studied to unravel the role of peribiliary vascular plexus at the basis of biliary defects. Biliary complications after liver transplantation, including non-anastomotic strictures (NAS), represent a major cause for re-transplantation, with a reported incidence varying between 5% and 30%; therefore, their pathogenesis would be crucial to individuate prognostic and clinical target. Thus, our results would clarify whether the pathogenesis of NAS is associated with BTSC niche and whether peribiliary vascular plexus could participate in this process. The correlation of histological study with clinical, radiological, and surgical parameters will unravel whether a proper activation of PBG niche both in terms of degree and mature fate commitment would be necessary for regenerating surface epithelium defects due to transplantation procedure.
Therefore, the results coming from objectives #2 would be crucial to understand anatomy and physiology of HPC and PBG stem cell niches, signals within the niche, and their involvement in biliary disease with regard to clinical correlations. In general, data obtained from this part of the project will be useful to clarify the pathogenesis of cholangiopathies fibrosis and the possible therapeutic approaches.
The objective #3 will clarify whether in adult human duodenum remains a population of cells with endoderm progenitor-like features and with the capability of differentiate to human hepatocytes.
The plasticity of duodenal cell of differentiating towards mature liver cells will be tested by injection into mice with experimentally induced NAFLD. This will allow to understand if injected cells engraft and have the capability to correct the liver disease by repopulation of liver parenchyma.
This part of the project will also optimize the procedures to isolate these cells from adult human organs. Human duodenal cells would represent a readily available autologous cell source and could be more easily translatable into clinical programs compared to reprogrammed cells.