Abstract
I
Cell-based therapies have been explored to treat human liver disease [5]. Recently, liver specialists at King's College Hospital successfully pioneered peritoneal cavity infusion of alginate-encapsulated hepatocytes. This was opted for as the recipient liver and kidneys were severely damaged, and therefore transplantation was not an option. As a result of the treatment, the patient's liver began to regenerate, providing proof of concept in juveniles. The use of this approach in adult liver disease poses a number of challenges, and is undergoing further development. Adult liver support has also been provided by extracorporeal liver devices and gained clinical support [6]. Most recently, exciting studies have highlighted the essential role of the macrophage in liver remodeling and regeneration post-injury [7,8]. These studies represent a major advance in the field that could lead to the development of immune matched and less invasive procedures to treat human liver disease in the future.
In addition to the clinic, hepatocytes provide useful human laboratory models. Prevention is of course the best way of reducing the morbidity and mortality associated with human disease. Therefore, the development and application of simple cell-based models, which accurately reflect human physiology in vivo is an important objective [9]. Such models will not only deliver a better understanding behind the disease process, but in the future promise the identification of biomarkers and medicines, leading to more effective medical intervention [10]. Developing reliable liver models in a dish is therefore an important objective [11].
In order for cell-based therapies to succeed in the clinic, approaches must employ technology that can be scaled to meet demand. Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) are examples of scalable stem cell populations, and when coupled with an efficient differentiation procedure, offer an unlimited supply of human soma. hESCs are derived from the inner mass of grade B blastocysts, which are unsuitable for human implantation. iPSCs are derived from human soma by inducing pluripotency using defined sets of transcription factors [12,13]. iPSCs display similar properties to hESCs, with the important advantage of being immune matched. This promises in the future that recipients may require little or no immunosuppression post-transplant. There has been a huge effort within the field to deliver pluripotent stem cell (PSC) scale-up in bioreactors. Culture systems have been developed that permit long-term expansion of PSCs in suspension culture [14]. Whilst exciting, other studies have demonstrated the variability of PSC lines to such approaches, highlighting the requirement for a number of lines to be tested using new protocols [15,16]. More recently studies have focused on large-scale and rapid expansion of compliant PSC lines. Encouragingly, the use of suspension cultures has delivered scale-up of hESCs and iPSCs to clinically relevant numbers in a relatively short time span [17]. This is a very exciting development as PSC populations have been shown to efficiently differentiate into many cell types, including hepatocytes, offering great potential in the quest for renewable cell based therapies [9].
The article “Generation of functional hepatocyte-like cells from human pluripotent stem cells in a scalable suspension culture” published in this month's Stem Cells and Development does exactly that and makes a fantastic contribution to the field [18]. The authors developed a system for mass manufacture of stem cell-derived hepatocytes in cell numbers that would be useful for clinical application. PSCs were adapted from adherent feeder-free cultures to suspension culture. Following this, definitive endoderm was specified and displayed equivalence to well-established two-dimensional adherent procedures. Subsequently, stem cell-derived hepatocyte differentiation was achieved using a two-step process. During hepatocyte differentiation, two different spheroid populations were produced, cystic and dense. The cystic structures displayed a high level of mesodermal gene expression, which was in contrast to hepatic gene expression detected in dense spheroids. The dense spheroids were selected and characterized in detail. iPSCs differentiated in this manner yielded of ∼55% albumin-positive cells and this was consistent with hESC yields. The differentiated cell populations displayed multiple hepatocyte functions, including glycogen storage, low-density lipoprotein uptake, indocyanine green uptake, cytochrome p450 activity, albumin secretion, and production of urea. Following specification, stem cell-derived hepatocytes were enriched and transplanted into a murine liver injury model. Encouragingly, PSC-derived hepatocytes transplantation led to a significant increase in the survival rate and a reduction in liver damage markers. Cell engraftment was estimated at ∼3%–4% and was focused around pericentral and periportal veins. Importantly, the authors detected no tumor formation in the liver or other major organs at 15 weeks post transplantation.
These studies provide an exciting development in the field, promising in future, that PSC populations and their derivatives can be scaled and manufactured at reasonable cost. This offers future possibilities that immune-matched cell-based therapies could be used clinically. Additionally, such approaches could permit the scale-up of stem cell populations, where mutated genes have been corrected, prior to differentiation and transplantation [19]. While these studies have shown great promise, there is a requirement to develop serum-free scale-up processes, which are generally applicable to PSC lines with minimal batch-to-batch variation. Additionally, it is essential to characterize the safety [20] and efficiency of stem cell derivative hepatocyte engraftment in other in vivo models. Importantly, in models of liver damage, consideration should also be given to the state of the tissue niche before cell transplantation, to ensure efficient cell engraftment and long-term stability [7,8].