Polymer Supported Directed Differentiation Reveals a Unique Gene Signature Which Predicts Stable Hepatocyte Performance

The ability to generate renewable sources of human soma, from defined genetic backgrounds, has enormous potential for modern medicine, with immediate impacts being the delivery of tailor-made models human tissue ‘in a dish’, with the delivery of cell-based therapies for degenerative disorders likely to follow. Theoretically, pluripotent stem cells (PSCs) can give rise to all somatic cell types found in the human body,[1] with their self-renewal and differentiation properties offering the prospect of generating unlimited quantities of human cells for biomedical application. Our particular interest lies in the delivery of human hepatocyte like cells (HLCs) derived from PSCs. Numerous hepatocyte differentiation protocols have been established using two- and three-dimensional cell culture,[2] and encouragingly, the stem cell derived HLCs produced exhibit many typical hepatocyte characteristics.[3-15] While these attributes are promising, the HLCs produced are immature in status, reminiscent of fetal or neo-natal hepatocytes. [16] One major obstacle to stem cell derived HLC maturation has been cellular instability in culture, similar to their primary human hepatocyte counterparts. Notably, the undefined or xenobiotic nature of cell culture components in both systems contribute to variable cell performance, making it difficult to unravel the complexity behind liver cell differentiation and dedifferentiation.
Therefore, the development of highly defined cell based systems is required if the true potential of stem cell derived HLCs is to be realised. Such systems should be simple to use, scalable and highly defined, and capable of delivering a “product“ with predictable performance and "shelf life“. There have been a number of approaches developed, including the use of differential cytokine or chemical combinations, three dimensional cell aggregation, and perfused devices, to mimic the liver niche and architecture.[17-20] While these approaches marked significant progress, their complexity and/or undefined nature has limited large scale deployment of the technology. In order to tackle this issue, we hereby report on the use of a tunable biopolymer substrate in conjunction with a serum free differentation procedure with research and GMP grade PSCs. Importantly, those stem derived populations displayed robust and predictable performance in cell culture which was hallmarked by gene expression of matrix metalloproteinase 13 (MMP13), delta catenin (CTNND2) and thrombospondin 2 (THBS2).