The generation of induced hepatic stem cells by direct conversion and its applications in cell-based therapy for liver disease treatment

Abstract

The liver is a central functional organ in the body. Even though the liver is made of remarkably regenerative tissue, chronic liver damage and inflammation induce loss of normal hepatic functions and lead to liver fibrosis and finally to cirrhosis of the injured liver by extracellular matrix remodeling. Liver transplantation is the only effective therapy for patients with end-stage liver fibrosis. However, the shortage of organ donation and limited primary hepatocyte propagation capacity remains a challenge for treating liver injury. Alternatively, stem cell-based therapy may be considered for liver treatment. Previous studies have demonstrated the generation of hepatocyte and hepatic stem cells from pluripotent stem cells (PSCs) by differentiation and direct conversion methods. PSCs are an essential cell source for regenerative medicine. PSCs, including embryonic stem cells and induced pluripotent stem cells, possess the ability to self-renew and the potential to differentiate into a variety of cell types derived from three germ layers. With the increasing importance of PSCs for cell-based therapy, the risk of tumorigenesis after transplantation due to residual undifferentiated PSCs within the differentiated progenies remains a challenge for clinical applications. Direct conversion bypassing the pluripotent state has been described in previous reports as an attractive method of acquiring hepatocytes for cell-based therapy. The limited proliferation of directly induced hepatic cells, however, has hampered its uses in cell-based therapy. The purpose of the present study was 1) to discover a novel human pluripotency sensor for the real-time screening of human PSCs (hPSCs) and study the mechanism of SHI5 to identify and isolate hPSCs; 2) to generate induced hepatic stem cells (iHepSCs) by direct conversion and explore their self-renewal and bipotency with the capacity to differentiate into functional hepatocytes and cholangiocytes; and 3) to study possible mechanisms of liver regeneration using iHepSCs in the CCl4-induced liver fibrosis model. This thesis/dissertation consists of the following: Chapter Ⅰ is a general introduction related to an overview of the liver and liver disease, strategy of hepatic cells generation for liver disease treatment, and an introduction of diversity-oriented fluorescent livery. Chapter Ⅱ explains the direct monitoring of live human pluripotent stem cells by a highly selective pluripotency sensor. Chapter Ⅲ demonstrate that Oct4 and Hnf4α-induced hepatic stem cells ameliorate chronic liver injury in the liver fibrosis model. In conclusion, I present a novel pluripotency sensor to overcome safety issue by real-time monitoring hPSCs for hPSC-based therapy and demonstrate a new strategy for acquiring a sufficient number of iHepSCs that may be an attractive cell source for liver disease treatment.