345 Cell culture media that are less proliferative and more homeostatic improve differentiation, responsiveness and co-culture of keratinocytes, fibroblasts and melanocytes in 2D and 3D culture

Abstract

Combinations of highly proliferative cell culture additives such as serum (FCS), pituitary extract (BPE), insulin and EGF/FGF-family members are routinely used to generate rapid in vitro proliferation of a variety of skin cell types. Highly concentrated proliferative signals such as these are ideal for amplifying cells, however they are poorly suited to many other situations where very different cell behaviours are required. We have investigated a range of less proliferative, more homeostatic media for their suitability when inducing differentiation and co-culturing keratinocytes, fibroblasts and melanocytes. Differentiation was improved in all three cell types, with 2x better barrier (3D keratinocytes), 2.5x more melanin (2D melanocytes), and 5x more ECM (2D fibroblasts). Homeostatic media were also found to increase responsiveness of melanocytes and fibroblasts to external stimuli such as tyrosinase inhibitors and vitamin C treatment respectively. Variants of standard proliferation media were also found to deliver balanced 2D co-culture of keratinocytes and fibroblasts, with each cell type delivering similar proliferation rates when seeded at equal density on standard tissue culture plastic. Finally, a fibroblast medium tailored for ECM production was also found to be ideally suited to the fast and simple establishment of full-thickness skin models with fully natural dermal matrix, thereby avoiding the handling challenges and variability associated with traditional collagen scaffolds. In conclusion, less proliferative, more homeostatic culture media were found to deliver a range of improved cell behaviours that complement the single rapid-growth behaviour created by standard proliferation media.