Since the first human hepatocyte transplants (HTx) in 1992, clinical studies have clearly established proof of principle for this therapy as a treatment for patients with acquired or inherited liver disease. Although major accomplishments have been made, there are still some specific limitations to this technology, which, if overcome, could greatly enhance the efficacy and implementation of this therapy. Here, we describe what in our view are the most significant obstacles to the clinical application of HTx and review the solutions currently proposed. The obstacles of significance include the limited number and quality of liver tissues as a cell source, the lack of clinical grade reagents, quality control evaluation of hepatocytes prior to transplantation, hypothermic storage of cells prior to transplantation, preconditioning treatments to enhance engraftment and proliferation of donor cells, tracking or monitoring cells after transplantation, and the optimal immunosuppression protocols for transplant recipients.

Farnesoid X receptor (FXR) is the master regulator of bile acid (BA) homeostasis because it controls BA synthesis, influx, efflux, and detoxification in the gut/liver axis. Deregulation of BA homeostasis has been linked to hepatocellular carcinoma (HCC), and spontaneous hepatocarcinogenesis has been observed in FXR-null mice. This dreaded liver neoplasm has been associated with both FXR gene deletion and BA-mediated metabolic abnormalities after inactivation of FXR transcriptional activity. In the present study, we addressed the hypothesis that intestinal selective FXR reactivation would be sufficient to restore the fibroblast growth factor 15 (FGF15)/ cholesterol-7alpha-hydroxylase (Cyp7a1) enterohepatic axis and eventually provide protection against HCC. To this end, we generated FXR-null mice with re-expression of constitutively active FXR in enterocytes (FXR 2/2 iVP16FXR) and corresponding control mice (FXR 2/2 iVP16). In FXR-null mice, intestinal selective FXR reactivation normalized BA enterohepatic circulation along with up-regulation of intestinal FXR transcriptome and reduction of hepatic BA synthesis. At 16 months of age, intestinal FXR reactivation protected FXR-null mice from spontaneous HCC development that occurred in otherwise FXR-null mice. Activation of intestinal FXR conferred hepatoprotection by restoring hepatic homeostasis, limiting cellular proliferation through reduced cyclinD1 expression, decreasing hepatic inflammation and fibrosis (decreased signal transducer and activator of transcription 3 activation and curtailed collagen deposition). Conclusion: Intestinal FXR is sufficient to restore BA homeostasis through the FGF15 axis and prevent progression of liver damage to HCC even in the absence of hepatic FXR. Intestinal-selective FXR modulators could stand as potential therapeutic intervention to prevent this devastating hepatic malignancy, even if carrying a somatic FXR mutation. (HEPATOLOGY 2015;61:161-170) See Editorial on Page 21 H epatocellular carcinoma (HCC) is the fifthmost prevalent type of cancer and the secondleading cause of cancer-related death, with over Abbreviations: AKR1B7, aldo-keto reductase 1b7; ALDH1B1, aldehyde dehydrogenase 1 family member B1; ALT, alanine aminotransferase; ANG1, angiogenin 1; ANOVA, analysis of variance; AST, aspartate aminotransferase; BA, bile acid; CA, cholic acid; CCND1 and E1, cyclinD1 and E1; CLD, chronic liver disease; CYP3A11, cytochrome P450 isoform 3A11; CYP7A1, cholesterol-7alpha-hydroxylase; ERK1/2, extracellular signal-regulated protein kinases 1 and 2; FGF15, fibroblast growth factor 15; FGFR4, FGF receptor 4; FRS2, FGF receptor substrate 2; FXR, farnesoid X receptor; GSTM1, glutathione S-transferase mu1; IBABP, ileal bile acid-binding protein; JNK, c-Jun (NH2)-terminal kinase; HCC, hepatocellular carcinoma; KO, knockout; IFN-g, interferon gamma; IHC, immunohistochemistry; IL-6, interleukin-6; b-MCA, beta-muricholic acid; MRP, multidrug resistance-associated protein; NF-jB, nuclear factor kappa B; NRF2, nuclear factor erythroid 2-r...

Differentiation of stem cells to hepatocyte-like cells (HLCs) holds great promise for basic research, drug and toxicological investigations, and clinical applications. There are currently no protocols for the production of HLCs from stem cells, such as embryonic stem cells or induced pluripotent stem cells, that produce fully mature hepatocytes with a wide range of mature hepatic functions. This report describes a standard method to assess the maturation of stem cell-derived HLCs with a moderately high-throughput format, by analysing liver gene expression by quantitative RT-qPCR. This method also provides a robust data set of the expression of 62 genes expressed in normal liver, generated from 17 fetal and 25 mature human livers, so that investigators can quickly and easily compare the expression of these genes in their stem cell-derived HLCs with the values obtained in authentic fetal and mature human liver. The simple methods described in this study will provide a quick and accurate assessment of the efficacy of a differentiation protocol and will help guide the optimization of differentiation conditions.

Different cell types can be isolated from human placental tissues, and some have been reported to retain phenotypic plasticity and characteristics that make them a promising source of cells for regenerative medicine. Among these are human amnion epithelial cells (hAECs). Adoption of current good manufacturing practices (cGMP) and enhanced quality control is essential when isolating hAECs in order to deliver a safe and effective cellular product for clinical purposes. This unit describes a detailed protocol for selective isolation of hAECs from human term placenta with little to no contamination by other cell types. A method for characterizing the heterogeneity of the hAEC suspension is also provided. The resulting cell product will be useful for clinical as well as basic research applications. © 2016 by John Wiley & Sons, Inc.

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