Human hepatic stem cells from fetal and postnatal donors.

Human hepatic stem cells (hHpSCs), which are pluripotent precursors of hepatoblasts and thence of hepatocytic and biliary epithelia, are located in ductal plates in fetal livers and in Canals of Hering in adult livers. They can be isolated by immunoselection for epithelial cell adhesion molecule-positive (EpCAM+) cells, and they constitute approximately 0.5-2.5% of liver parenchyma of all donor ages. The self-renewal capacity of hHpSCs is indicated by phenotypic stability after expansion for >150 population doublings in a serum-free, defined medium and with a doubling time of approximately 36 h. Survival and proliferation of hHpSCs require paracrine signaling by hepatic stellate cells and/or angioblasts that coisolate with them. The hHpSCs are approximately 9 microm in diameter, express cytokeratins 8, 18, and 19, CD133/1, telomerase, CD44H, claudin 3, and albumin (weakly). They are negative for alpha-fetoprotein (AFP), intercellular adhesion molecule (ICAM) 1, and for markers of adult liver cells (cytochrome P450s), hemopoietic cells (CD45), and mesenchymal cells (vascular endothelial growth factor receptor and desmin). If transferred to STO feeders, hHpSCs give rise to hepatoblasts, which are recognizable by cordlike colony morphology and up-regulation of AFP, P4503A7, and ICAM1. Transplantation of freshly isolated EpCAM+ cells or of hHpSCs expanded in culture into NOD/SCID mice results in mature liver tissue expressing human-specific proteins. The hHpSCs are candidates for liver cell therapies.

Differentiated human adipose-derived stem cells exhibit hepatogenic capability in vitro and in vivo.

The availability of suitable human livers for transplantation falls short of the number of potential patients. In addition, the availability of primary human hepatocytes for cell-therapy and drug development applications is significantly limited; less than 700 livers per year are available for such studies. However, the majority of these organs cannot be utilized due to pathological infections (e.g., HepB, HepC, or HIV) or excessive levels of steatosis. Thus, the number of cells needed for cell therapy applications far exceeds the number of cells available from donated livers. The ability to implant progenitor cell populations that can form liver tissue in situ, or can be differentiated in vitro would be a major advance in current cell-based therapies. In addition, and importantly for this application, the ability to utilize a non-hepatic progenitor cell to mimic hepatocytes in vitro would enable the scale-up production of cells for bioartifical liver assist devices, cell-therapy and drug discovery applications. We demonstrate the feasibility of inducing adipose-derived stromal (ASC) cells to express several features of human hepatocytes such as glycogen storage and expression of liver specific genes. Importantly, we also show that undifferentiated ASCs and ASC-derived hepatic cells engraft robustly into the liver in a mouse model of toxic injury. These data indicate a significant potential for the use of undifferentiated ASCs and ASC-derived hepatic cells as novel and valuable products for cell therapy.

Cytokine profile of human adipose-derived stem cells: expression of angiogenic, hematopoietic, and pro-inflammatory factors.

Adipose tissue serves as a source of adipokines and cytokines with both local and systemic actions in health and disease. In this study, we examine the hypothesis that multipotent human adipose-derived stem cells (ASCs), capable of differentiating along the adipocyte, chondrocyte, and osteoblast pathways, contribute to adipose tissue-derived cytokine secretion. Following exposure to basic fibroblast growth factor (bFGF) or epidermal growth factor (EGF), the ASCs significantly increase their secretion of hepatocyte growth factor (HGF), a cytokine implicated in hematopoiesis, vasculogenesis, and mammary epithelial duct formation. Ascorbic acid synergizes with these inductive factors, further increasing HGF levels. Following exposure to lipopolysaccharide, ASCs increase their secretion of both hematopoietic (granulocyte/monocyte, granulocyte, and macrophage colony stimulating factors, interleukin 7) and proinflammatory (interleukins 6, 8, and 11, tumor necrosis factor alpha) cytokines based on ELISA and RT-PCR. In co-cultures established with umbilical cord blood-derived CD34(+) cells, the ASCs support long-term hematopoiesis in vitro. Furthermore, in short-term 12-day co-cultures, the ASC maintain and expand the numbers of both myeloid and lymphoid progenitors. These observations are consistent with the functionality of the secreted cytokines and confirm recent reports by other laboratories concerning the hematopoietic supportive capability of ASCs. We conclude that the ASCs display cytokine secretory properties similar to those reported for bone marrow-derived mesenchymal stem cells (MSCs).