Generation of human hepatocytes by stem cell technology: definition of the hepatocyte

Abstract

Since 1999, numerous articles have reported the generation of hepatocytes from different types of extrahepatic stem or precursor cells. This opens exciting new possibilities for pharmacology and toxicology, as well as for cell therapy. Hepatocyte marker expression, including albumin, cytokeratin 18, c-met, alpha-fetoprotein and cytochrome P450 3A4 and -2B6, has been observed after transplantation of different types of human stem cells into the liver of laboratory animals or in vitro after incubation with cytokines. These intriguing observations have prompted scientists to classify stem cell-derived cell populations as hepatocytes. However, this conclusion may be premature. It has been shown that factors of the liver microenvironment can induce expression of a limited number of hepatocyte marker genes in nonhepatic cell types. To conclude on the grounds of a limited number of markers that these cells are true hepatocytes is not indicated. In this case one should carefully evaluate crucial hepatocyte-defining enzymatic properties. The present article: i) reviews studies describing the fate of extrahepatic human stem and precursor cells in livers of laboratory animals, including the possibility of cell fusion; and ii) critically discusses the phenotype of stem cells after application of various differentiation protocols aimed at generating human hepatocytes. In addition, the necessary criteria needed for defining a true hepatocyte are suggested. Establishing the necessary properties for stem cell-derived hepatocytes is timely and reasonable, and thus avoids further misleading semantic confusion. Finally, it is essential to understand that the definition of a bona fide hepatocyte should not be limited to qualitative assays, such as reverse transcriptase polymerase chain reaction and immunohistochemistry, but has to include a quantitative analysis of enzymatic activities, which allows direct comparison with primary hepatocytes. Although the stem cell-derived-hepatocyte does not yet exist there is a good chance that this aim may be achieved in the future.

Fate of extrahepatic human stem and precursor cells after transplantation into mouse livers

“…The first evidence that hematopoietic stem cells might be capable of differentiating into hepatocytes in rodents came from Petersen et al 3 Similar results were obtained in further studies using different animal models and purified cell types for transplantation (for a review, see Hengstler et al 1 ). Later, it was recognized that cell fusion and not transdifferentiation was responsible in the FAH Ϫ/Ϫ mouse model.…”

“…1,2 At first glance, this appears to provide exciting new opportunities for cell therapy, as some types of stem cells proliferate efficiently in vitro and therefore may help to generate a larger supply of human hepatocytes or precursor cells for transplantation. On the other hand, some studies presenting far-reaching conclusions with respect to the capacity of stem cell therapy have not yet been reproduced or may have been interpreted in an overly optimistic manner.…”

“…The intriguing results obtained in rodents stimulated many groups to study the fate of human stem and precursor cells in livers of experimental animals (Supplemental Table 1; for a review, see Hengstler et al 1 ). Recipients were mostly immunodeficient severe combined immunodeficiency (SCID) or nonobese diabetic (NOD)/SCID mice.…”

Highlight report: perspectives in stem cell research—unbiased quantification of the similarity between in vitro generated and primary hepatocytes

“…However, it is discussed controversially to which degree stem cell-derived cells resemble primary cells. In the field of stem cell-derived hepatocyte-like cells, some authors reported that generation of 'functional,' 'high fidelity' hepatocytes from stem and precursor cells is already possible (Medine et al 2013;Szkolnicka et al 2014;Huang et al 2014), while others present a more critical point of view Hengstler et al 2005;Brulport et al 2007).…”

Alcohol-induced liver injury: how a small molecule overwhelms one of the cell types with the best regeneration capacity of the human body

“…In addition in larger animals and humans, the regeneration capacity of the liver is excellent. Another experimental system to study the liver regeneration is that of mice deWcient in fumarylaetoacetate hydrolase causing a permanent deterioration of hepatocytes and Wnally death of the animal (review: Hengstler et al 2005). Hepatocytes transplanted into these livers colonize and rescue the entire organ (review: Nussler et al 2006).…”