R ESEARCH ARTICLE 10.2217/17460751.2.3.289 © 2007 Future Medicine Ltd ISSN 1746-0751 Regenerative M ed. (2007) 2(3), 289–300 289 part of Transplantation of 3D scaffolds seeded with human embryonic stem cells: biological features of surrogate tissue and teratoma-forming potential Justin G Lees 1 , Sue Anne Lim 2 , Tristan Croll 2 , Georgia Williams 1 , Sylvia Lui 1 , Justin Cooper-White 2 , Leon R McQuade 1 , Bagyalakshmi Mathiyalagan 1 & Bernard E Tuch 1† † Author for correspondence 1 University of New South Wales, Diabetes Transplant Unit, Prince of Wales Hospital, Sydney, Randwick, NSW 2031, Australia Tel.: +61 293 824 814; Fax: +61 293 824 826; E-mail: b.tuch@unsw.edu.au 2 University of Queensland, Tissue Engineering and Microfluidics Laboratory, Division of Chemical Engineering and the Australian Institute for Bioengineering and Nanotechnology, Brisbane, Australia Keywords: 3D cell culturing, human embryonic stem cells, scaffolds, surrogate tissue, teratomas Aim: To generate complex surrogate tissue by transplanting 3D scaffolds seeded with human embryonic stem cells (hESCs) between the liver lobules of severe combined immunodeficient (SCID) mice and to assess the teratoma-forming potential. M aterials & methods: 3D poly-(lactic-co-glycolic acid) (PLGA) scaffolds coated with laminin were seeded with hESCs and then transplanted between the liver lobules of SCID mice. After a period of in vivo differentiation, the scaffolds were retrieved and analyzed using reverse transcription polymerase chain reaction, immunofluorescent staining and scanning electron microscopy. Results: A proportion of the hESCs within the scaffolds differentiated into cells that produced proteins characteristic of specific tissues, including endoderm and pancreatic markers glucogon-like peptide-1 receptor, islet amyloid polypeptide and Insulin. Markers of hepatic and neuronal lineages were also investigated. Major matrix proteins abundant in multiple tissue types, including collagen I, laminin and collagen IV, were found to be profuse within the scaffold pores. Transplantation of the seeded scaffolds between liver lobules also resulted in extensive vascularization both from host blood vessel incursion and the differentiation of hESCs into endothelial progenitor cells. An investigation of teratoma- forming potential demonstrated that transplantation of 3D scaffolds seeded with hESCs will, under certain conditions, lead to the growth of teratomas. Discussion: Transplantation of 3D scaffolds seeded with hESCs between liver lobules resulted in the development of surrogate tissue containing cells that produced proteins representing the pancreatic, hepatic and neuronal lineages, the assembly of an extracellular matrix structure and the formation of a vasculature. hESCs seeded within 3D scaffolds and transplanted into SCID mice were capable of forming teratomas. However, the formation and progression of teratoma growth is shown to be dependant on both the site of transplantation and the treatment of cells prior to transplantation. Regenerative medicine involves the develop- ment of surrogate tissue for the replacement of diseased or damaged tissues. Self renewal of human embryonic stem cells (hESCs) in a pluripotent state provides a unique source of cells for prospective therapies. hESCs differen- tiated in a monolayer can be induced to exhibit characteristics of specific cell phenotypes, such as definitive endoderm progenitors and insu- lin-producing β-cells [1,2]. The differentiation strategies used to achieve such outcomes are based on the ontology of precursor cells and involve exposure to factors that are known to play key roles in tissue development. T he responsiveness of hESCs to this type of signal- ing indicates that efficient differentiation strat- egies must provide hESCs with factors that are encountered by target tissue progenitors during in vivo development [3]. Another important factor that is characteris- tic of in vivo tissue development is the forma- tion of elaborate 3D structures. The structure of a tissue will, in most circumstances, incorpo- rate a complex extracellular matrix (ECM) and an intricate vasculature. It has been demon- strated that the efficient function of multiple cell types, including islet hormone-producing cells, is dependent on matrix-producing and endothelial cells that provide a 3D support structure and sufficient vascularization [4–6]. It has also been demonstrated that islets do not function efficiently postisolation without resto- ration of an effective matrix structure [7]. T here- fore, the 3D interactions between numerous cell types plays an important role in the func- tion of tissue and may also be required for the efficient production of surrogate tissue struc- tures derived from hESCs. Indeed, it has been For reprint orders, please contact: reprints@futuremedicine.com