Down-regulation of RASA1 Is Associated with Poor Prognosis in Human Hepatocellular Carcinoma.

BACKGROUND/AIM: RASA1 (p120RasGAP), encodes Ras GTPase-activating protein 1 and, is a potent tumor suppressor gene that is frequently inactivated in several human cancer types. However, its precise role in hepatocellular carcinoma (HCC) has been blurred. MATERIALS AND METHODS: We hypothesized that RASA1 plays a crucial role in tumor pathogenesis and progression of HCC. RASA1 expression levels were analyzed in 226 cases of HCC by immunohistochemistry. RESULTS: It was found that 38.68% (41/106) of the high-grade HCC samples and 54.17% (65/120) of the low-grade HCC samples expressed RASA1 protein. The difference between RASA1 expression in high-grade and low-grade HCC was statistically significant (p=0.02). Additionally, RASA1 high expression was inversely associated with larger tumor size (p<0.001). Although RASA1 is known as a tumor suppressor, its role in overall survival (OS) in HCC is unclear. Kaplan-Meier survival analysis showed that patients with low level of RASA1 expression correlated with a significantly poorer survival compared to those with high level of RASA1 expression. CONCLUSION: These data support that RASA1 could serve as an independent prognostic marker for HCC patients.

Paracrine signals from mesenchymal cell populations govern the expansion and differentiation of human hepatic stem cells to adult liver fates.

UNLABELLED: The differentiation of embryonic or determined stem cell populations into adult liver fates under known conditions yields cells with some adult-specific genes but not others, aberrant regulation of one or more genes, and variations in the results from experiment to experiment. We tested the hypothesis that sets of signals produced by freshly isolated, lineage-dependent mesenchymal cell populations would yield greater efficiency and reproducibility in driving the differentiation of human hepatic stem cells (hHpSCs) into adult liver fates. The subpopulations of liver-derived mesenchymal cells, purified by immunoselection technologies, included (1) angioblasts, (2) mature endothelia, (3) hepatic stellate cell precursors, (4) mature stellate cells (pericytes), and (5) myofibroblasts. Freshly immunoselected cells of each of these subpopulations were established in primary cultures under wholly defined (serum-free) conditions that we developed for short-term cultures and were used as feeders with hHpSCs. Feeders of angioblasts yielded self-replication, stellate cell precursors caused lineage restriction to hepatoblasts, mature endothelia produced differentiation into hepatocytes, and mature stellate cells and/or myofibroblasts resulted in differentiation into cholangiocytes. Paracrine signals produced by the different feeders were identified by biochemical, immunohistochemical, and quantitative reverse-transcription polymerase chain reaction analyses, and then those signals were used to replace the feeders in monolayer and three-dimensional cultures to elicit the desired biological responses from hHpSCs. The defined paracrine signals were proved to be able to yield reproducible responses from hHpSCs and to permit differentiation into fully mature and functional parenchymal cells. CONCLUSION: Paracrine signals from defined mesenchymal cell populations are important for the regulation of stem cell populations into specific adult fates; this finding is important for basic and clinical research as well as industrial investigations.

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.

Identification and characterization of vitamin A-storing cells in fetal liver: implications for functional importance of hepatic stellate cells in liver development and hematopoiesis.

Hepatic stellate cells (HpSTCs) are major regulators of hepatic fibrogenesis in adults. However, their early development in fetal liver is largely unknown. To characterize fetal HpSTCs in the liver, in which hepatic development and hematopoiesis occur in parallel, we determined the phenotypic characteristics of HpSTCs from rat fetal livers, using a strategy focused on vitamin A. Storage of vitamin A in the cytoplasm is a unique characteristic of HpSTCs, permitting identification of them by vitamin A-specific autofluorescence (vA+) when excited with UV light using flow cytometry. A characteristic vA+ cell population was identified in liver as early as 13 days post coitum; it had a surface phenotype of RT1A- intercellular adhesion molecule (ICAM)-1+ vascular cell adhesion molecule (VCAM)-1+ beta3-integrin+. Although nonspecific autofluorescent cells were found with the antigenic profile of RT1A- ICAM-1+ VCAM-1+, they were beta3-integrin- and proved to be hepatoblasts, bipotent hepatic parenchymal progenitors. In addition to expression of classic HpSTC markers, the vA+ cells were able to proliferate continuously in a serum-free hormonally defined medium containing leukemia inhibitory factor, which was found to be a key factor for their replication. These results demonstrated that the vA+ cells are fetal HpSTCs with extensive proliferative activity. Furthermore, the vA+ cells strongly express hepatocyte growth factor, stromal-derived factor-1alpha, and Hlx (homeobox transcription factor), indicating that they play important roles for hepatic development and hematopoiesis. The abilities to isolate and expand fetal HpSTCs enable further investigation into their roles in early liver development and facilitate identification of possibly novel signals of potential relevance for liver diseases.