Cellular and molecular mechanisms of liver regeneration
Greenhalgh, Stephen Nicholas
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Improved understanding of how the liver regenerates would be of great value, particularly given the dearth of therapies for end-stage liver disease. Currently, the only effective treatment for total liver failure is transplantation. Such an invasive, costly and specialised intervention is unable to address the enormous global impact from diseases of the liver. Ironically, the liver has the greatest regenerative potential of any organ in the mammalian body. However, this capacity for repair is overwhelmed in the face of massive or repeated injury. Understanding the key factors driving or inhibiting successful liver regeneration offers the potential for novel, targeted therapies to promote regeneration of a patient’s own liver. Animal models are widely used when studying complex, dynamic, multicellular processes such as liver injury and regeneration. Continued progress in transgenic modification of mice, combined with ongoing advances in microscopy techniques, means that the opportunity now exists to observe labelled cells, and subcellular structures, in real time and in vivo, with previously unobtainable resolution and fidelity. Not only does this afford the opportunity for novel insights into both normal physiology and the response to injury or disease, it can vastly expand the amount of biologically relevant information that can be obtained from each experimental animal. Hence, it is possible to advance scientific knowledge and reduce experimental animal use simultaneously. This thesis examines the role of αv integrins in liver regeneration. Integrins are expressed on the surface of cells and can perform a range of functions, including signalling and extracellular matrix adhesion. The most well-characterised role for αv integrins is activation of transforming growth factor beta, a molecule which has been shown to inhibit hepatocyte proliferation and liver regeneration. Partial hepatectomy was used as an experimental model of liver injury and regeneration. It was performed in mice, in which one or more αv integrins had been genetically depleted from specific cell types in the liver, namely hepatocytes, hepatic stellate cells or liver sinusoidal endothelial cells. These investigations revealed that depletion of integrin αvβ8 from hepatocytes led to increased hepatocyte proliferation and accelerated liver regeneration. The possible mechanisms through which hepatocyte integrin αvβ8 may exert its braking effect on liver regeneration following injury were also explored. In parallel, a novel experimental system to permit intravital multiphoton microscopy of the regenerating liver following partial hepatectomy in mice was developed and validated. Intravital imaging of mouse liver was performed with a range of cellular labels, combined with a fluorescent cell cycle reporter and label-free imaging modalities. This demonstrated the enormous potential of the system to study the dynamics of hepatocytes and non-parenchymal cells in the regenerative niche, reconstruct the sinusoidal vascular network in three dimensions during angiogenesis, and measure sinusoidal blood flow and parenchymal lipid deposition. Advances in experimental animal models such as this drive forward our understanding of the cellular and molecular mechanisms of liver regeneration whilst refining and reducing experimental animal use. Novel insights into the process of liver regeneration will permit the development of innovative therapeutic strategies to allow this remarkable organ to heal itself even in the face of massive or sustained insult.