Liver organoid systems to identify metabolic signalling functions underlying liver regeneration and enhance drug discovery.

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The liver performs many functions vital to our health, including protein synthesis, digestion and drug metabolism/detoxification. Strikingly and in contrast to other internal organs, when damaged the liver is capable of multiple rounds of regeneration. Recently, 3D cell aggregates, known as organoids, have been described that can model liver regeneration in vitro. How regenerative signals integrate intra- and extra-cellular cues, including the energy balance of the cell, to guide adult stem cells to exit quiescence, proliferate and differentiate is not clear. This project will use organoids derived from mouse and human adult liver to elucidate the metabolic signalling functions underlying liver regeneration. Advances in metabolomics have revealed multiple systems in which metabolic substrates and byproducts regulate stem cell fate via a balance of glycolysis and oxidative phosphorylation. Organoids provide tangible systems in which to assess the activity and effect of specific metabolic signalling pathways during differentiation in a controlled environment separate from the influence of whole-body metabolism or other signalling cues. This project aims to; 1) Gain full metabolic control of liver organoid culture using fully chemically-defined media and a synthetic hydrogel as 3D scaffold. 2) Characterise metabolic state during regeneration using enzymatic assays and imaging of organoids with a genetically encoded FRET-based metabolic flux sensor. 3) Investigate the functional relevance of metabolic state during regeneration through metabolic perturbations. Perturbations will be achieved by alterations to oxygen and nutrient availability, administration of metabolic inhibitors, and through genetic manipulation of rate-limiting glycolytic enzymes. Greater insight into liver regeneration is a key interest for basic biology and ultimately may help derive therapies for patients suffering from chronic liver injury. Importantly, understanding the metabolic requirements of liver progenitors will lead to protocols to better recapitulate functional liver models in vitro for therapeutic purposes such as personalised medicine, drug discovery and toxicity studies.