Technology

Our ambition is to provide cells with conditions which allow them to develop functionality which closely mimics that seen in the intact organism.

To achieve this, we have developed a system which obviates the need for scaffolds, gels or any type of additives which might perturb gene expression away from its natural baseline equilibrium. Furthermore we have very significantly reduced contact with plastics and reduced shear forces. Apart from this, it is possible to use the culture media normally used for cell culture (but where we would suggest and reducing glucose levels to those closer to those seen in animals).

 

The CelVivo system creates an environment which promotes the functionality of large 3D tissue mimetic structures, whether they are spheroids, organoids, acini and other aggregates.

The technology CelVivo use for culturing cells is based on the clinostat principle also (incorrectly) referred to as microgravity system.

A clinostat, (ClinoStar), and a rotating bioreactor (ClinoReactor) keep the cells suspended by causing the gravitation forces to act on the cells from all sides (thus technically omnidirectional normogravity and not microgravity).

This  approach has the advantages that the spheroids or organoids formed are exposed to very low shear forces, have good gas and nutrient exchange.

Spheroids formed using microwell plates and are very similar in size (after 21 days in culture (which would amplify any differences) they still only show a 21% variation in protein content. This allows them to grow up to about 1mm without showing necrosis or apoptosis in the core (although some form of inactivity does develop there).

    Figure 1
    1) cells; 2) culture media; 3) culture vessel

    The advantages of using clinostats for culturing cells are:

    Large spheroids are created with high uniformity in size
    Excellent for co-cultures and tumor studies
    The cells experience very low shear stress
    Active diffusion which allow good nutrient exchange.
    The function, architecture and ultrastructure of the spheroid mimics that see in vivo after 3 weeks

    Spheroids grown using clinostat rotated bioreactors from human C3A cells (an immortal hepatocellular carcinomacellline) have been shown to need 18 days to reach maturity (as determined by their growth rate, viability and their production of urea, cholesterol and ATP). After this time they are metabolically stable for at least 24 days more; grow slowly (a doubling time of >20 days); produce physiological levels of urea, cholesterol and ATP; exhibit stable gene expression (for selected liver relevant genes); and can post-translationally modify proteins in a manner which mirrors those seen in vivo.

    Studies with 6 common drugs (acetaminophen, amiodarone, diclofenac, metformin, phenformin, and valproic acid) have shown that they are more predictive of lethally toxic plasma levels in vivo than published studies using primary human hepatocytes. Shotgun proteomics has revealed that the gain of in-vivo-like physiological properties is due to [SJF2] modulation of the levels of hundreds of cellular proteins

    The spheroids grown using the CelVivo clinostats for culturing cells are:

    • Toxicology studies -both acute and chronic, (single dose or repeated-dose)
    • Efficacy studies
    • Cancer research
    • Regenerative medicine
    • Drug discovery