Cytochrome P450 Expression of Cultured Rat Small Hepatocytes after Long-Term Cryopreservation

Abstract

ABSTRACT:Small hepatocytes (SHs) are hepatic progenitor cells that can be cryopreserved for a long time. After thawing, the cells can proliferate and, when treated with Matrigel, they can differentiate into mature hepatocytes (MHs). In this study, we investigated whether cryopreserved SHs could express cytochromes P450 (P450s), whether P450 expression was induced by appropriate inducers, and whether P450 activities were measurable.

Thyroid Hormone Is Necessary for Expression of Constitutive Androstane Receptor in Rat Hepatocytes

“…As previously reported (Ooe et al, 2006), SH colonies consisting of 30 to 50 cells were observed at day 14 after plating. To collect SHs, the colonies were detached from dishes and immediately replated (subculture) or cryopreserved at Ϫ80°C until use.…”

“…Although, unlike in MHs, PB could not induce RXR␣ expression in the cryo-SHs, these results showed that low expression of CAR might be involved in the poor expression of CYP2B1. Comparing the previous (Miyamoto et al, 2005;Ooe et al, 2006) and present experiments, the difference of the experimental procedure was whether cells were cryopreserved. Thus, we compared the expression of major nuclear receptors involved in the regulation of P450 expression between sub-SHs and cryo-SHs.…”

“…A goat anti-CYP2B1 (Daiichi Pure Chemicals Co., Tokyo, Japan) antibody was used for immunoblots. The details of the method were described previously (Ooe et al, 2006).…”

“…On the other hand, we reported that SHs could be cryopreserved for more than 6 months and that, even after thawing, the cells could maintain the capability of the growth and the maturation . We also examined the expression of CYP1A1/2, CYP2E1, and CYP3A in thawed SHs even after the cells were cultured in a serum-free medium for more than 1 month (Ooe et al, 2006). However, CYP2B1 was not expressed in the culture even if hepatic maturation of SHs was induced by Matrigel and induction by phenobarbital was carried out (Ooe et al, 2006).…”

Selective proliferation of rat hepatocyte progenitor cells in serum-free culture

“…Manipulation of the growth and maturation of SHs is easy, and they can be cryopreserved 17 . Cryopreserved SHs can maintain the abilities of growth and maturation 18 . Maturated SHs in serum-free culture can be used for studies of hepatic drug metabolism, liver diseases and regenerative medicine.…”