Grant-in-Aid for Scientific Research (C)
|Research Institution||KYUSHU UNIVERSITY|
松下 琢 九大, 工学部, 助教授 (10209538)
IJIMA Hiroyuki KYUSHU UNIV., Chem.Eng., ASSISTANT PROFESSOR, 工学部, 講師 (10274515)
KIHARA Shin-ichi KYUSHU UNIV., Chem.Eng., RESEARCH ASSISTANT, 工学部, 助手 (30284524)
FUNATSU Kazumori KYUSHU UNIV., Chem.Eng., PROFESSOR, 工学部, 教授 (80037960)
|Project Fiscal Year
1997 – 1998
Completed(Fiscal Year 1998)
|Budget Amount *help
¥3,300,000 (Direct Cost : ¥3,300,000)
Fiscal Year 1998 : ¥1,100,000 (Direct Cost : ¥1,100,000)
Fiscal Year 1997 : ¥2,200,000 (Direct Cost : ¥2,200,000)
|Keywords||Drug Metabolism Simulator / Alternatives to Animal Testing / Polyurethane Foam / Multi-Channel PUF Packed-Bed / Hepatocyte / Spheroid / Acetaminophen / Lidocaine / 薬物代謝シミュレーター / 動物実験代替 / ポリウレタン発泡体 / 多細管PUF充填層 / 肝細胞 / スフェロイド / アセトアミノフェン / リドカイン|
We developed drug metabolism simulator by investigating the following items.
1.Establishment of a large amount of primary hepatocytes from large animals.
We developed a coII agenase perfusion system of a liver and filtrated method using stainless steel mesh (106 mum) to prepare the large amount of he patocy te s form large animals. 180 g hepatocytes were obtained by 10kg pig (300 g liver). This procedure took 2.5 hours, and the viability of the hepatocytes was about 60 %.
2.Establishment of rapid formation conditions of spherical hepatocytes tissue like organoid (spheroid).
It is takes 3-7 days culture to form hepatocytes spheroid in the pores of polyurethane foam (PUF). In this case, we originally male a hydrophilic PUF which was made by changing the composition of the ratio of polyol and isocyanate. Rat, dog and porcine hepatocytes were formed spheroid in about 1 day of culture. Furthermore, we found that autocrine subustances of hepatocytes affect the spheroid formation period.
ion of the biotransformation of the drugs by hepatocytes spheroid.
Lidocaine, acetamincphen aid 7-ethoxycoumarin were used as a model drugs, and evaluate the drug biotransformation activities of rat, dog and porcine hepatocytes spheroid. These biotransformation activities expressed the sane as the liver in vivo. Furthermore, these functions were maintained for two weeks of culture, but its were lost within two days of monolayer culture.
4.Evaluation of the induction ability of drug metabolism enzymes by spheroid.
We evaluate the response of hepatocytes spheroid by the induction of drug metabolism enzyme activity using 3-methylcholanthorene. Rat hepatocytes spheroid was induced the drug metabolism activity even in two weeks culture.
5.Evaluation of the drug metabolizing rate of hepatocytes spheroid using perfusion culture system.
We developed PUF/hepatocytes spheroid packed-bed type perfusion culture module. 4% of a rat whole liver cells were immobilized in this module, and evaluate the metabolizing rate of lidocaine, acetaminophen as a model drug. The metabolizing rate per a unit cell number was as same as the rate of rat whole liver perfusion. Less