Stem cells in liver disease

Abstract

Failure of the liver, the largest vital organ in the body, unequivocally results in death. Hepatic failure most commonly evolves over a period of several years as a result of chronic liver disease, most often viral hepatitis or alcoholic liver damage. In rarer cases, the organ shuts down within weeks or even days, in a clinical picture referred to as fulminant hepatic failure. Currently, orthotopic liver transplantation is the only effective measure that can avert mortality when liver function has become insufficient to sustain life. However, transplantation is hampered by a severe shortage in donor organs, and the procedure is costly and requires harmful, lifelong immunosuppressive treatment. Stem cell technologies may one day provide alternative treatment options for end-stage liver disease that could alleviate these problems. A selection of the potential future therapeutic modalities involving stem cells is investigated in this thesis. In particular, experimental treatment using mesenchymal stem cells (MSCs) - an intriguing stem cell type with proven therapeutic effects in models of myocardial infarction, stroke, kidney failure and several other diseases – is investigated. In this thesis, we show that MSCs can be employed to cure rats undergoing fulminant hepatic failure (D-galactosamine model) using different MSC-based treatment modalities. The most effective method involves hooking up the circulation of the animal to an MSC-filled bioreactor. This method is complex and may therefore not be appealing in the clinical setting. MSC lysates injected intravenously into rats undergoing liver failure had a similar therapeutic effect. In addition, a single bolus injection of exclusively the >3Kd fraction of the secreted molecules from MSCs resulted in significantly improved survival, representing a simple, off-the shelf treatment option that does not require injection of cellular, immunogenic material. In further studies, we demonstrate that cytokines, chemokines and growth factors secreted by the MSCs are involved in their therapeutic effects in liver disease. In the rat model of fulminant hepatic failure, and using in vitro studies with primary hepatocytes, we present evidence that molecules secreted by the MSCs: 1) stimulate hepatocyte regeneration, 2) inhibit liver cell death, and 3) dampen harmful immune responses. This last effect, the inhibiting effect on immune cell function and inflammation, is also observed in an in vitro model of liver fibrosis. Using neutralizing antibodies, we demonstrate that MSCs decrease pro-fibrogenic functions of activated stellate cells, the primary mediators of liver fibrosis/cirrhosis, through secreted interleukin-10, tumor necrosis factor-alpha and hepatocyte growth factor. The last chapters of the thesis are devoted to two different types of experimental stem cell applications in liver disease: 1) fetal stem/progenitor cells for tissue engineering applications, and 2) gene-knockout technologies (for which embryonic stem cells are often used) aimed at xenotransplantation of livers. In in vitro studies, 1) a method for reversible boosting of fetal liver stem/progenitor cell functions is described, and 2) the extent of reduction of immune rejection of porcine liver tissue by human and baboon serum is quantified. In conclusion, several types of stem cell technologies may have potential as future treatment modalities for human liver disease. In particular, MSCs reverse liver failure and increase survival in a fulminant hepatic failure model in the rat through pro-regenerative, anti-apoptotic and anti-inflammatory actions of MSC-secreted molecules. Further studies are needed to confirm the effects in different animal models of fulminant hepatic failure and to dissect the mechanisms involved before human trials may be attempted.