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New organs to help us Liver longer?

“Vascularized and functional human liver from an iPSC-derived organ bud transplant”  

The ex vivo manufacture of organs destined for use as transplants has gained much momentum following a recent paper in Nature, in which scientists from multiple centres in Japan have reported the generation of three-dimensional (3D) vascularised and functional human liver tissue from human induced pluripotent stem cells (hiPSCs).   Led by Professor Hideki Taniguchi at Yokohama City University, the team demonstrated that hiPSCs can produce liver (hepatic) cells which can self-organise into 3D cell clusters that, when transplanted into mice, are able to connect with host vessels and integrate within the host system, maturing into cells that are very similar to that of adult human liver tissue in their structure and function.1

The authors started by using standard protocols2 to generate hepatic endoderm from hiPSCs (iPSC-HEs), yielding cells which were positive for the hepatic marker HNF4A.   During normal development, stromal and endothelial cells provide essential cues for liver formation, so the team co-cultured iPSC-HEs with human umbilical vein endothelial cells (HUVECs) and human mesenchymal stem cells (MSCs) under two-dimensional (2D) conditions. Surprisingly, they found that this resulted in the self-assembly of 3D organoids resembling embryonic liver buds (iPSC-LBs) that had formed their own endothelial network surrounded by iPSC-HEs within just 2 days.   Microarray profiling revealed strong upregulation of FGF and BMP pathways in iPSC-HEs, and further experiments indicated an important previously uncharacterised role for stromal cell-dependent paracrine support in 3D liver bud formation.   However, the signature of iPSC-LBs indicated the retention of an early developmental stage.  

As haemodynamic stimulation is necessary for liver bud maturation during development, the authors then placed intact 3D liver organoids upon the brain of immunodeficient mice, taking advantage of the extensive blood supply existing at this site.   By 48 hours, Takebe et al. observed the development of HUVEC-derived blood vessels, which connected with the host circulation and went on to form a complex vascular network.   Hepatic cells within transplanted liver buds continued to proliferate for the first two months, and by the end of this period significant reorganisation and maturation had occurred within the organoid, in which hepatic cords fashioned from cuboidal shaped hepatocytes (iPSC-Heps) formed bile canaliculi with neighbouring iPSC-Heps, partially resembling the structure of adult liver tissue.   Organoids were also shown to survive at additional extrahepatic sites including under the kidney capsule and into the mesentery (the peritoneum connecting the small intestine to the dorsal abdominal wall), and demonstrated impressive albumin production at these sites from 10 days after transplantation.   Importantly, Takebe et al. demonstrate adult human liver-like protein production and drug metabolism in iPSC-derived liver cells, indicating some functionality.   The 3D iPSC-LBs microenvironment was found to enhance the maturation of iPSC-Heps over 2D approaches, enabling 3D cells to outperform 2D cells in liver function tests.   Immunocytochemical studies, however, confirmed the absence of bile ducts within organoids, which may indicate insufficient time for tissue maturation following transplantation or that additional cues are required for these final liver elements to form.   Nonetheless, the authors go on to demonstrate a very exciting result, that the mesenteric transplantation of hiPSC-derived liver tissue equivalent to 1 percent of the mouse hepatocyte mass was sufficient to rescue subacute liver failure in mice.  

The shortage of donor livers for transplantation to treat end-stage organ failure is a worldwide problem.   Autologous stem cell therapies require little or no immunosuppression and entail reduced ethical considerations compared with embryonic stem cell based therapy, however the generation of complex vascularised organ-like structures until now had not yet been achieved.   The exciting and promising results reported by Takebe et al. are the most promising thus far in the field of liver bioengineering, made even more attractive by the inherent capacity of the liver to integrate newly formed cells. The limitations posed by transplanting de novo hepatocytes into patients with liver disease on the long term survival and function of grafted tissue, however, remains an important issue for exploration.   Transplanting organoids into an extrahepatic microenvironment proffering the necessary bloody supply, as achieved in this work, is a promising alternative, allowing the release of bile into the circulation provided that the host liver retains some excretory function.   Prior attempts to recellularize denuded liver scaffolds have yielded limited results; however the use of 3D liver organoids as building blocks within such scaffolds retaining the vascular structure may enhance the success of this approach and allow the bioengineering of a more complete organ.   This is not the first report demonstrating the capability of stem cells to ‘self-organise’, in fact self-morphogenesis in vitro has recently attracted much media attention (for example the self-formation of optic cups containing stratified neural retina from human ESCs), however the fabrication of other ‘life-saving’ organs until now has proven more difficult.   This work floods new light on the potential of liver bioengineering and has no doubt injected much excitement into the field.   While it is clear that much remains to be done before hiPSC-derived liver tissue can be transplanted into humans for therapeutic purposes, this work consolidates the enormous potential of iPSCs for cellular liver therapy and narrows the distance between on-going research at the bench and patients awaiting treatment in the clinic.

 

Reference

1. T. Takebe, K. et al. (2013) Vascularized and functional human liver from an iPSC-derived organ bud transplant. Nature 499(7459):481-4. Epub 2013 Jul 3.

2. Si-Tayeb, K. et al. (2010) Highly efficient generation of human hepatocyte-like cells from induced pluripotent stem cells. Hepatology 51:297–305. 

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Transplantable liver organoids made from only three ingredients, Cell Stem Cell

Japanese scientists use stem cells to create human liver, Japan Daily Press

The Quest for an Artificial Liver, MIT news Magazine

Perspectives on human micro-liver-like structures made from iPS cells, Knoepfler Lab Stem Cell Blog

Livers Created from Stem Cells, The Scientist

Japanese scientists grow working mini-livers for mice from stem cells, The Independent

Tiny stem cell livers grown in laboratory, BBC Health News

Scientists Fabricate Rudimentary Human Livers, The New York Times

Miniature human liver grown in mice, Nature News

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Scientists use stem cells to grow human livers in mice, The Guardian

 

From Nature by Takebe et al.

Stem Cell Correspondent Carla Mellough reports on those studies appearing in current journals that are destined to make an impact on stem cell research and clinical studies.