In drug discovery research, proper evaluation of incorporation, metabolism and excretion of a drug or the like in the hepatic tissue of a living body is extremely important. Further, for simply carrying out large-scale drug screening, or from an ethical point of view, an in vitro test method has been demanded. As a means for such a test, cultured cells are used, and their culture is preferably carried out under conditions reflecting those in a living body as much as possible. For example, as a culture system for hepatocytes, one which can excellently reproduce the proper polarity of the hepatic tissue of a living body, wherein, for example, the cell membrane is clearly differentiated between the blood vessel side and the bile canaliculus membrane side, is demanded
For example, Patent Document 1 describes that it is thought that, by binding a binding protein or an adhesion protein to a microscale substrate and culturing hepatocytes thereon, formation of a bile canaliculus is promoted. Further, Patent Document 2 describes that, by culturing small hepatocytes on a polycarbonate porous sheet covered with collagen, a bile canaliculus-like structure was formed on Day 30 of the culture.
In the above-described three-dimensional culture methods, preparation of hepatocytes having the polarity from the primary cultured cells takes several weeks to several months.
On the other hand, in cases where the collagen gel sandwich method (Non-patent Document 1: LeCluyse et al., Am J Physiol Cell Physiol, 1994, vol. 266, pp. 1764-1774) is used, formation of a bile canaliculus and the bile component excretion activity begins to be detected about 3 or 4 days after deposition of a collagen gel. However, even in the cases where the collagen gel sandwich method is used, several days are required before the bile component excretion activity is obtained. Further, metabolites cannot be continuously analyzed in such cases since a bile duct having an outlet, as is the case in a living body, is required for the continuous analysis. There is also a problem in that the number of bile canaliculi is too small and the activity is too low, to be used in drug screening.
Further, in Non-patent Document 2 (Tissue Engineering vol.13 Number 8 2007 2105-2117) and Non-patent Document 3 (Acta Biomaterialia 5 2009 613-620), it is shown that, when hepatocytes are to be cultured on gas-permeable PDMS (polydimethylsiloxane), adhesion of the cells can be maintained for 3 days by coating the surface of the PDMS with PEM or by forming thin pillars having a diameter of 1 to 3 μm. However, the adhesion cannot be maintained for a long time and hence those cultured cells are problematic in view of stable use in a test. Further, it has been generally thought that PDMS is not suitable for adherent culture of hepatocytes.