We recently derived mouse expanded potential stem cells (EPSCs) from individual blastomeres by inhibiting the critical molecular pathways that predispose their differentiation 1. EPSCs had enriched molecular signatures of blastomeres and possessed the developmental potency for all embryonic and extraembryonic cell lineages. Here, we report the derivation of porcine EPSCs, which express key pluripotency genes, are genetically stable, permit genome editing, differentiate to derivatives of the three germ layers in chimeras, and produce primordial germ celllike cells in vitro. Under similar conditions, human ESCs and iPSCs can be converted, or somatic cells directly reprogrammed, to EPSCs that display the molecular and functional attributes reminiscent of porcine EPSCs. Significantly, trophoblast stem cell-like cells can be generated from both human and porcine EPSCs. Our pathwayinhibition paradigm thus opens a new avenue for generating mammalian pluripotent stem cells, and EPSCs present an unique cellular platform for translational research in biotechnology and regenerative medicine.

Hepatic stellate cell activation is a main feature of liver fibrogenesis. We have previously shown that phagocytosis of apoptotic bodies by stellate cells induces procollagen ␣1 (I) and transforming growth factor beta (TGF-␤) expression in vitro. Here we have further investigated the downstream effects of phagocytosis by studying NADPH oxidase activation and its link to procollagen ␣1 (I) and TGF-␤1 expression in an immortalized human stellate cell line and in several models of liver fibrosis. Phagocytosis of apoptotic bodies in LX-1 cells significantly increased superoxide production both in the extracellular and intracellular milieus. By confocal microscopy of LX-1 cells, increased intracellular reactive oxygen species (ROS) were detected in the cells with intracellular apoptotic bodies, and immunohistochemistry documented translocation of the NADPH oxidase p47phox subunit to the membrane. NADPH oxidase activation resulted in upregulation of procollagen ␣1 (I); in contrast, TGF-␤1 expression was independent of NADPH oxidase activation. This was also confirmed by using siRNA to inhibit TGF-␤1 production. In addition, with EM studies we showed that phagocytosis of apoptotic bodies by stellate cells occurs in vivo. In conclusion, these data provide a mechanistic link between phagocytosis of apoptotic bodies, production of oxidative radicals, and the activation of hepatic stellate cells.

Hepatic fibrosis is a wound-healing process that occurs when the liver is injured chronically. Hepatic stellate cells (HSC) are responsible for the excess production of extracellular matrix (ECM) components. The activation of HSC, a key issue in the pathogenesis of hepatic fibrosis, is mediated by various cytokines and reactive oxygen species released from the damaged hepatocytes and activated Kupffer cells. Therefore, inhibition of HSC activation and its related subsequent events, such as increased production of ECM components and enhanced proliferation, are crucial goals for intervention in the hepatic fibrogenesis cascade. This is especially true when the etiology is unknown or there is no established therapy for the cause of the chronic injury. This review explores the rationale for choosing HSC as a target for the pharmacological, molecular, and other novel therapeutics for hepatic fibrosis. One focus of this review is the inhibition of two cytokines, transforming growth factor-beta and platelet-derived growth factor, which are important in hepatic fibrogenesis. A number of new agents, such as Chinese herbal recipes and herbal extracts, silymarin, S-adenosyl-L-methionine, polyenylphosphatidylcholine, and pentoxifylline are also discussed.

Background & Aims Our previous studies showed that CD133, EpCAM and aldehyde dehydrogenase (ALDH) are useful markers to identify cancer stem cells (CSCs) in hepatocellular carcinoma (HCC) tissues. The present study aims to evaluate chemosensitivity and invasion capability of HCC based on CSC marker profiles, and to explore underlying molecular mechanisms. Methods Hepatoma cell lines were separated into subpopulations according to CD133, EpCAM and ALDH expression profiles. Epithelial mesenchymal transition (EMT) and hedgehog (Hh) signaling were examined to identify their links with chemoresistance and aggressive invasion. Results Well-differentiated cell lines were positive for CD133+/ALDHhigh and CD133+/EpCAM+ at 1.5–15% and 2.3–8.3%; whereas, poorly-differentiated cells were almost all negative for these markers. FACS-enriched CD133+/ALDHhigh and CD133+/EpCAM+ Hep3B and Huh-7 cells formed more spheroids in vitro. CD133−/ALDHlow HLE cells were more resistant to cisplatin, doxorubicin or sorafenib than their positive counterparts. CD133−/EpCAM− Huh-7 cells or CD133−/ALDH− HLE cells exhibited a higher invasion rate than their positive counterparts. HLE and HLF cells acquired EMT in double negative subpopulations. Hh activity in Huh-7 CD133−/EpCAM− cells was higher than in their positive counterparts, and the inhibition of Hh activity by cyclopamine resulted in reduced cell proliferation. Conclusions Well-differentiated CD133+/ALDHhigh or CD133+/EpCAM+ cells appear to be a CSC/initiating subpopulation; whereas, in poorly-differentiated hepatoma cells, EMT and enhanced hedgehog signaling activity may be responsible for their chemoresistance and invasion. These findings underscore the significance of EMT and enhanced Hh signaling in liver cancer stem or initiating cells.

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