Mechanobiology of Liver Fibrosis: Role of Hepatocytes in Fibrosis Progression

Abstract

Liver fibrosis is characterized by liver scarring which disrupts the liver architecture and impairs hepatic function. Current diagnostic tools reveal that the stiffness of the liver tissue increases as one advances through fibrosis, cirrhosis, and hepatocellular carcinoma. We investigated how changes in the mechanical microenvironment of the liver under different fibrotic conditions influence the biomechanical response of the liver cells. Among the different liver cells, it is believed that Hepatic Stellate Cells (HSCs) are the main perpetrators of liver fibrosis. On the other hand, hepatocytes have been mostly thought of as victims or bystanders. We studied the more active role of hepatocytes in liver fibrosis, as an instigator (that activates the HSCs to initiate and exacerbate fibrosis) or an accomplice (that directly causes changes to the ECM). We also studied liver fibrosis in the context of Hepatitis C infection – a major contributor to chronic liver injury leading to cirrhosis and liver cancer. To achieve the aforementioned research objectives, an in vitro system was established to simulate the mechanical stiffness of normal, fibrotic, and cirrhotic liver conditions. This was done by fabricating collagen-coated polyacrylamide gels of the relevant physiological stiffnesses, and culturing hepatocytes and HSCs on them. Both monoculture and co-culture conditions were implemented alongside the in vitro gel system to understand cell-cell and cell-ECM interactions during fibrosis progression. Hepatocytes expressing Hepatitis C viral proteins were also cultured and their biomechanical response to changes in substrate stiffness was also investigated and compared against that of the normal hepatocytes. Using the leads from our experimental results, we also developed a framework to simulate the interplay among the hepatocytes, HSCs, and their surrounding ECM under normal conditions and conditions of HCV infection and fibrosis. We also modelled cell death and division and analyzed how they changed the total energy of the system over many iterations. From the experimental and simulation work of this thesis, it can be concluded that both hepatocytes (normal and expressing viral proteins) and HSCs show significant biomechanical responses to changes in the stiffness of the substrate. It was also evident that the hepatocytes expressing the viral proteins exhibited a more sensitive response to changes in substrate stiffness when compared to normal hepatocytes. The cells also behaved differently in co-culture conditions as compared to monoculture conditions. It was also evident that hepatocytes do play a more active role in liver fibrosis but more along the lines of an instigator of HSCs that instigated the HSCs to exacerbate the conditions of fibrosis.