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Hello, I am going to be isolating equine hepatocytes and am trying to decide what culture technique would be best. I plan on using the hepatocytes for drug metabolism studies for in vivo extrapolation. I plan on cryopreserving some and using others fresh. There is a lot of literature discussing various techniqies, e.g. monolayer, gel suspension, sandwich, spherical 3D, microliver perfused 3D system.
I would like to know from the PK/PD community what the current opinions are for hepatocyte culture and if there are and references or protocols available that would be greatly appreciated as well.
From my readings it seems that the perfused 3D culture system is the best approach, however not coming from a biomedical engineering background it is not feasible for me to attempt this myself. If anyone knows how I could go about purchasing one please let me know or I am very willing to work in collaboration with a lab that has already developed this system.
Thank you in advance,
-- Kristin Grimsrud
DVM/PhD Graduate Candidate Pharmacokinetics, Pharmacodynamics & Pharmacometrics (P3)
Pharmacology/Toxicology Graduate Group
School of Veterinary Medicine University of California, Davis
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if your purpose is a metabolic profiling study of a new chemical entity, you could use your fresh hepatocytes in suspension, that is the easiest way, and it allows to obtain reliable results. You could test 2 or 3 drug concentrations and 2 or 3 time points, up to 4hr, and then choose the most representative sample.
If your compound has issues of poor solubility, you could indeed plate your hepatocytes in monolayer on a collagene-coated well plate. This system allows you to get also the 24hr time point (and even further), that, based on my experience, in some cases could be different, either qualitatively or quantitatively, from the 4hr.
It is true, indeed, that the other systems you mentioned, as the sandwich culture or the spherical 3D, represent a more physiological model, where the hepatocytes 3D conformation is more similar to the physiological one. Anyway, the suspension and the monolayer culture are reliable, as well. Fabio Carini
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