Hepatic stellate cells contribute to progenitor cells and liver regeneration

2014-12-01 00:01:10

The Journal of Clinical Investigation; 2014;124(12):5503–5515; DOI:10.1172/JCI74119

Claus Kordes, Iris Sawitza, Silke Götze, Diran Herebian and Dieter Häussinger


Retinoid-storing hepatic stellate cells (HSCs) have recently been described as a liver-resident mesenchymal stem cell (MSC) population; however, it is not clear whether these cells contribute to liver regeneration or serve as a progenitor cell population with hepatobiliary characteristics. Here, we purified HSCs with retinoid-dependent fluorescence-activated cell sorting from eGFP-expressing rats and transplanted these GFP+ HSCs into wild-type (WT) rats that had undergone partial hepatectomy in the presence of 2-acetylaminofluorene (2AAF) or retrorsine, both of which are injury models that favor stem cell–based liver repair. Transplanted HSCs contributed to liver regeneration in host animals by forming mesenchymal tissue, progenitor cells, hepatocytes, and cholangiocytes and elevated direct bilirubin levels in blood sera of GUNN rats, indicating recovery from the hepatic bilirubin–handling defect in these animals. Transplanted HSCs engrafted within the bone marrow (BM) of host animals, and HSC-derived cells were isolated from BM and successfully retransplanted into new hosts with injured liver. Cultured HSCs transiently adopted an expression profile similar to that of progenitor cells during differentiation into bile acid–synthesizing and –transporting hepatocytes, suggesting that stellate cells represent a source of liver progenitor cells. This concept connects seemingly contradictory studies that favor either progenitor cells or MSCs as important players in stem cell–based liver regeneration.


Hepatic stellate cells (HSCs) are characterized by exceptionally high amounts of retinoids in their quiescent state. These are mainly stored as retinyl palmitate in membrane-coated lipid vesicles and exhibit characteristic retinoid fluorescence after excitation by UV light. HSCs are primarily known as collagen-producing cells in the liver, which are responsible for fibrogenesis in chronic liver diseases. Despite intensive research for decades, HSCs have remained enigmatic. For instance, stellate cells express molecular markers of different germ layers, which impede a clear statement about their origin. The presence of glial fibrillary acidic protein (GFAP) and many other neuroectodermal proteins in stellate cells led to the view that they may derive from the neural crest. This concept has been questioned by microscopic as well as fate-mapping analysis in the developing liver, which indicated that stellate cells apparently originate from mesenchymal cells located in the subendothelial space of the septum transversum. This view is supported by the expression of mesodermal marker proteins such as desmin and &alpha -smooth muscle actin (&alpha -SMA or actin &alpha 2/ACTA2), especially in activated HSCs.

Until recently, HSCs were mainly studied with regard to their fibrogenic potential in chronic disease, whereas their identity and function in normal liver received little attention. Recent data suggest that stellate cells represent liver-resident mesenchymal stem cells (MSCs) due to their MSC-related expression profile, their potential to differentiate into adipocytes or osteocytes, and their supportive effects on extramedullary hematopoiesis. In line with this, stellate cells can originate from the bone marrow (BM), where MSCs were initially discovered, and reside in the liver close to endothelial cells, as is the case for MSCs in other organs. Quiescent stellate cells are typically located between sinusoidal endothelial cells and hepatocytes in the space of Disse, which has characteristics of stem cell niches. Moreover, activated stellate cells can develop into hepatocyte-like cells in vitro, and fate-mapping experiments using Gfap and Acta2 promoters indicate that they contribute to liver regeneration in vivo. In contrast to this, lineage-tracing experiments with lecithin retinol acyltransferase (LRAT) and mesoderm posterior 1 homolog (MESP1) failed to demonstrate a differentiation of HSCs into epithelial cells during regeneration of the injured mouse liver. Interestingly, LRAT and MESP1, which are suggested to be specifically expressed by HSCs or their precursors, are expressed by embryonic and adult stem cells from mice and humans. Owing to these opposing lineage-tracing studies, we performed cell transplantation experiments with HSCs in the present study to determine their contribution to liver regeneration. Transplanted stellate cells from the pancreas were recently reported to participate in liver repair through differentiation into epithelial cell lineages in a stem cell–based in vivo liver regeneration model. Also, MSCs from the BM and adipose tissue were shown to differentiate into liver parenchymal cells. Tissue-specific engraftment (homing), trafficking to sites of organ injury, and participation in tissue repair are important properties of stem cells that can be analyzed by cell transplantation experiments. Stem cells can be repeatedly transplanted into different organisms, and their retransplantation capacity is considered to represent one of their key features. Cell transplantation studies are therefore required to further validate stem cell characteristics of HSCs. In view of the generation of hepatocytes from HSCs, the question arises whether established liver progenitor cells, which are called oval cells in rodents, could represent an intermediate stage during hepatic differentiation of stellate cells or other MSCs. Liver progenitor cells are small cells that transiently appear after severe liver injury to restore liver mass when the proliferation of hepatocytes is impaired. These progenitor cells are identified by the expression of epithelial cell adhesion molecule (EPCAM), keratin 19 (K19), and &alpha -fetoprotein and protrude from the portal field into the injured liver tissue by forming duct-like structures (ductular reaction). Liver progenitor cells are reported to originate from stem cells with epithelial characteristics, but they also show features of mesodermal cells, such as the expression of vimentin. The present study addresses whether progenitor cells can derive from MSC populations such as hepatic stellate cells with mesodermal characteristics. The data show that transplanted HSCs can home to the injured liver and contribute to tissue regeneration by developing into putative progenitor cells and epithelial as well as mesenchymal tissues. In this process, progenitor cells seem to represent an intermediate stage of differentiating HSCs and other MSC populations.

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