Liver Regeneration after Resection and Transplantation: Mechanisms and Therapeutic Strategies

Since the Greek myth about Prometheus’ torture and the first scientific documentation of the phenomenon of liver regeneration in the 19th century, scientists have extensively investigated this intriguing process in an attempt to unravel its mystery. Numerous molecules and pathways involved in regeneration of the liver have been revealed, however the exact underlying mechanisms are still not fully elucidated. Meanwhile, the extensive regenerative capacity of the liver has been used to benefit patients with (end-stage) liver disease, as it enables oncologic liver resections and living-donor cq. split liver transplantation. However, several factors like a patient’s age, life style, nutritional status, disease condition, degree of injury and medication, but probably also genetic predisposition, can interfere with and limit the process of regeneration, resulting in impaired liver function and compromised homeostasis. Better understanding of the factors influencing and regulating liver regeneration after injury, contributes to the investigation and development of potential therapeutic strategies to prevent liver dysfunction and promote regeneration, thereby decreasing subsequent patient morbidity and mortality. The first part of this thesis highlights molecular mechanisms and functional pathways involved in liver regeneration after resection and transplantation. Differences in gene expression profiles between living liver donors with successful and incomplete regeneration of their remnant liver suggest a possible inhibition or delay in initiation of regenerative pathways in the incompletely regenerating livers. Similar, pathways and networks involved in the development of early allograft dysfunction (EAD) show downregulation of metabolic capabilities and upregulation of pro-inflammatory molecules. We defined a diagnostic gene expression signature to detect liver grafts prone to develop EAD. Furthermore, we report that inhibition of the regulatory protein mTOR by the immunosuppressant rapamycin severely impairs liver regeneration and show that this process can be partly reversed by exogenous growth factor treatment. The second part provides evidence for the presence of mesenchymal stromal/stem cells (MSCs) in the adult human liver. These cells have phenotypic and functional characteristics similar to bone marrow MSCs and migrate from liver grafts at time of transplantation. MSC cultures were evaluated for the presence of aberrant cells, showing that spontaneous malignant transformation is rare and only occurs after long-term culture. Finally, the effects of MSC-secreted factors on liver regeneration after surgical resection with or without ischemia and reperfusion injury show that treatment with these factors is a promising new strategy to modulate and accelerate liver regeneration after surgical injury

Mesenchymal Stromal Cell-Derived Factors Promote Tissue Repair in a Small-for-Size Ischemic Liver Model but Do Not Protect against Early Effects of Ischemia and Reperfusion Injury

Loss of liver mass and ischemia/reperfusion injury (IRI) are major contributors to postresectional liver failure and small-for-size syndrome. Mesenchymal stromal cell-(MSC-) secreted factors are described to stimulate regeneration after partial hepatectomy. This study investigates if liver-derived MSC-secreted factors also promote liver regeneration after resection in the presence of IRI. C57BL/6 mice underwent IRI of 70% of their liver mass, alone or combined with 50% partial hepatectomy (PH). Mice were treated with MSC-conditioned medium (MSC-CM) or unconditioned medium (UM) and sacrificed after 6 or 24 hours (IRI group) or after 48 hours (IRI + PH group). Blood and liver tissue were analyzed for tissue injury, hepatocyte proliferation, and gene expression. In the IRI alone model, serum ALT and AST levels, hepatic tissue damage, and inflammatory cytokine gene expression showed no significant differences between both treatment groups. In the IRI + PH model, significant reduction in hepatic tissue damage as well as a significant increase in hepatocyte proliferation was observed after MSC-CM treatment. Conclusion. Mesenchymal stromal cell-derived factors promote tissue regeneration of small-for-size livers exposed to ischemic conditions but do not protect against early ischemia and reperfusion injury itself. MSC-derived factors therefore represent a promising treatment strategy for small-for-size syndrome and postresectional liver failure

Support of hepatic regeneration by trophic factors from liver-derived mesenchymal stromal/stem cells

Mesenchymal stromal/stem cells (MSCs) have multilineage differentiation potential and as such are known to promote regeneration in response to tissue injury. However, accumulating evidence indicates that the regenerative capacity of MSCs is not via transdifferentiation but mediated by their production of trophic and other factors that promote endogenous regeneration pathways of the tissue cells. In this chapter, we provide a detailed description on how to obtain trophic factors secreted by cultured MSCs and how they can be used in small animal models. More specific, in vivo models to study the paracrine effects of MSCs on regeneration of the liver after surgical resection and/or ischemia and reperfusion injury are described

Detection of spontaneous tumorigenic transformation during culture expansion of human mesenchymal stromal cells

Human mesenchymal stem/stromal cells (MSCs) have been explored in a number of clinical trials as a possible method of treating various diseases. However, the effect of long-term cell expansion in vitro on physiological function and genetic stability is still poorly understood. In this study, MSC cultures derived from bone marrow and liver were evaluated for the presence of aberrant cells following long-term expansion. In 46 independent cultures, four batches of transformed MSCs (TMCs) were found, which were all beyond the culture period of five weeks. These aberrant cells were first identified based on the appearance of abnormal cytology and the acquirement of growth advantage. Despite common MSC markers being diminished or absent, TMCs remain highly susceptible to lysis by allogenic natural killer (NK) cells. When transplanted into immunodeficient mice, TMCs formed sarcoma-like tumors, whereas parental MSCs did not form tumors in mice. Using a combination of high-resolution genome-wide DNA array and short-tandem repeat profiling, we confirmed the origin of TMCs and excluded the possibility of human cell line contamination. Additional genomic duplication and deletions were observed in TMCs, which may be associated with the transformation event. Using gene and microRNA expression arrays, a number of genes were identified that were differentially expressed between TMCs and their normal parental counterparts, which may potentially serve as biomarkers to screen cultures for evidence of early transformation events. In conclusion, the spontaneous transformation of MSCs resulting in tumorigenesis is rare and occurs after relatively long-term (beyond five weeks) culture. However, as an added safety measure, cultures of MSCs can potentially be screened based on a novel gene expression signature