Traditional cell culture

Traditional tissue culture is based on growing mammalian cells that have been selected for growth under the conditions provided in the culture. These cultures are based on growing cells in a monolayer on treated polystyrene (tissue culture plastic, TCP) in the presence of nutrient-rich basal media (e.g. DMEM, RPMI 1640) that has been supplemented with animal sera. The cells that survive under these conditions are malignant. They are transformed stem/progenitor cells that are immortal and have one or more genetic modifications that obviate their ability to lineage restrict to mature cells. This allows them to be cultured indefinitely, something that cannot be done with normal cells. However, since by their very nature they are not normal, using them to understand in vivo cell behavior has limits. This limitation is especially apparent when trying to create functional tissues ex vivo, such as a liver. Note that even if primary cells are harvested properly, they can rarely be cultured on TCP in traditional media and retain their function.  

Making a functional tissue ex vivo

To recreate complex, functional tissue ex vivo, it is readily apparent that providing only soluble signals in the form of media and growing cells in a monolayer on TCP is insufficient. At a minimum, the tissue substrata that supports the cells (the extracellular matrix, ECM) must also be included in the culture conditions. Not only does the ECM provide the proper structure for cell growth, it is also required for telling the cells whether or not to migrate, differentiate or initiate apoptosis. However, to make a complicated system even more complex, soluble and matrix signals are still only part of the picture. The oxygen levels that the cells are exposed to are also important – this is especially obvious for stem cells that seem to prefer hypoxic conditions. And finally, the nature of the physical forces (e.g. stress, strain) that cells are exposed to are also critical for their function.

Cell culture matrices

Significant work remains before we are able to fully recreate most functional tissues ex vivo. However, culturing cells with a cell culture matrix and using the proper media allows for a much closer emulation of in vivo conditions than can be achieved with cultures on TCP. Additionally, these matrices, when used in conjunction with bioreactors, have the capacity to recreate the in vivo environment ex vivo. Cell culture matrices can be anything from fractured polystyrene (that provide a non-uniform, varied surface) to electrospun fibers (that imitate collagen fibrils in size and whose chemistry can be tailored to the application) to ECM proteins (that coat TCP and provide different attachment sites for cells) to naturally-derived basement membrane (that provide a compliant surface rich in ECM proteins and growth factors) to semi-synthetic or synthetic hydrogels (that provide compliant surfaces and can be customized with the relevant ECM proteins).

What is Extracel?

Extracel is composed of thiol-modified hyaluronic acid (Glycosil™) and gelatin (Gelin-S™) that are co-crosslinked with polyethylene glycol diacrylate (Extralink™) to form a semi-synthetic ECM.  In Extracel, Gelin-S provides the attachment signals in the form of denatured collagen. However, denatured collagen can provide inappropriate signals to the cells being cultivated, especially stem cells. Therefore, in place of Gelin-S, ECM proteins like laminin, collagen, vitronectin, or fibronectin can be non-covalently incorporated into our collagen-free HyStem hydrogels. Finally, growth factors are also retained within the Extracel and HyStem hydrogels.

Why Extracel™ ?

Extracel provides a stripped down version of the ECM, giving scientists an excellent research tool to build their own cell-specific ECM by controlling the hydrogel composition and rigidity. In the absence of detailed information about specific matrix conditions, researchers can use Extracel alone to promote cell attachment and cultivate cells. Additionally, cells can either be encapsulated in Extracel to simulate a more 3-D like environment or they can be plated on the surface of a pre-formed hydrogel or sponge.