Investigating the differentiation of tissue structure using 3D cell technologies

Abstract

The capability for Embryonic Stem Cells (ESCs) to self-renew indefinitely in culture whilst maintaining their capacity to differentiate is of keen interest in research into developmental biology and, the pharmaceutical industry for novel regenerative and personal therapies. To apply the use of ESCs to these fields, their characterisation is a vital step towards ensuring differentiation capacity and, a simple model to test for putative stem cell differentiation capacity is in demand. Currently, the ‘gold standard’ test for a putative stem cell population and their differentiation capacity is via the teratoma assay, which involves injecting the cell population into mice, allowing for a teratoma to form. A stem cell population will form a teratoma consisting of ectodermal, endodermal and mesoderm tissues. Due to the complexity of differentiation that occurs when a teratoma is allowed to form, the development of an in vitro model to recapitulate all the factors occurring in vivo remains elusive. These complex tissue structures that can be generated from the teratoma that are unable to be seen in in vitro models, may be due to the constraints of the cell culture environment.

In this project, we used 3D cell technologies, namely Alvetex® Scaffolds, to overcome this constraint. Alvetex® provides a platform that allows cells to not be constricted to an x-y planar topography that is seen in conventional 2D culture, which can reduce the developmental potency of ESCs and thus their capacity to differentiate. We found that mES CGR8 cells that were cultured in 3D had greater differentiation capabilities that those cultured in 2D and, expressed antigens that were immunoreactive to ectodermal, endodermal and mesodermal lineage, the resounding factors that are necessary in a teratoma assay to deem a putative cell population to have stem cell properties.