1990 Fiscal Year Annual Research Report

培養肝細胞を用いた機能と増殖を調節する肝組織構築の生化学的研究

Research Project

Project/Area Number	02454156
Research Institution	The University of Tokushima
Principal Investigator	市原明徳島大学, 酵素科学研究センター, 教授 (40035374)
Co-Investigator(Kenkyū-buntansha)	冨田優美子徳島大学, 酵素科学研究センター, 助手 (00089913) 田中啓二徳島大学, 酵素科学研究センター, 助手 (10108871) 野田千征子徳島大学, 酵素科学研究センター, 助教授 (40035506)
Keywords	肝細胞 / 初代培養 / 細胞接触 / 細胞周期 / 肝再生 / 肝臓機能
Research Abstract	(1)肝細胞Spheroid及び単層培養による細胞周期調節 Primaria培養皿でラット肝細胞をinsulin.EGFを含むWilliams E(WE)培地で3日培養するとSpheroidを形成した。Spheroidでは増殖促進に十分な条件でもDNA合成はほとんどみられずAlbuminの強い発現がみられ典型的Go期にあると考えられた。Spheroidをcollagen塗布培養皿に移し培養すると徐々にSpheroidが崩れて単層を成形した。そして単層形成に従ってDNA合成の促進とAlbumin量の低下がみられた。更に培養すると分裂像がみられた。以上の結果は肝細胞の機能と増殖は細胞接触(或は細胞の形)によって強く相反的に調節されている事を示す。また単層培培養とSpheroid培養を相互に転換することにより細胞周期を調節できるようになり細胞周期を解析する為の有効なin vitro実験系が確立できた。 (2)肝細胞における脂肪酸合成酵素肝特異酵素のPyruvate Lactateによる誘導肝細胞を高密度でinsulin、Dexamethasone、T3を含むWE培地にPyruvate又はLactateを30mM添加して6日間培養すると細胞の維持が良くFatty acid synthetase、Glucoseー6ーPhosphate dehydrogenase、Malic enzyme Citrate cleavage enzyme等の脂肪酸合成系の酵素とGlucokinaze,Aldolase B等の肝特異酵素の誘導が著明となった。この事からATPの受生と肝細胞の機能発現は特異的に相関していることが示唆された。 (3)肝臓障害時の肝細胞増殖因子(HGF)の増加ラットに四塩化炭素を与えると一日後に肝HGF活性は20倍増加する。また肝硬変患者腹水中にも増加する。いづれも精製しその性質は吾々の発見したHGFと同じであることを確認した。

Research Products

(13 results)

All Other

All Publications (13 results)

[Publications] Akira ICHIHARA: "Regulation of protein and amino acid metabolisms in cultured hapatocytes" Nutrion:Protein and Amino Acids. 49-66 (1990)
[Publications] Chiseko NODA: "Organization and strucutre of 5'flanking region of the rat serine dehydratase gene" J.Biochem.108. 622-628 (1990)
[Publications] Chiseko NODA: "Developmental and growth related regulation of expression of serine dehydratase mRNA in liver" Biochem.Biophys.Res.Commun.168. 335-342 (1990)
[Publications] Keiji TANAKA: "Proteasomes (Multicatalytic proteinase complex) in eukaryotic cells" Cell Struct.Funct.15. 127-132 (1990)
[Publications] Chizuko Tsurumi: "cDNA cloning and sequencing for the import precursor of subunit B H^+ーATP synthase from rat mitochondria" Biochem.Biophys.Res.Commun.169. 136-142 (1990)
[Publications] Tomihiko HIGUCHI: "cDNA cloning and sequencing for the import precursor of coupling factor 6 in H^+ーATP synthase from rat liver mitochondria" Biochem.Biophys.Res.Commun.171. 1079-1086 (1990)
[Publications] Osamu ASAMI: "Purification and characterization of hepatocyte growth factor from injured liver of carbon tetrachloride treated rats" J.Biochem.109. 8-31 (1991)
[Publications] Shigeru YOSHIZAWA: "Inhibition of protein phosphatases by microcyctin and nodularin associated with hepatotoxicity" J.Cancer Clin Oncol.116. 609-614 (1990)
[Publications] Ichiro SHIMIZU: "Hepatocyte growth factor in ascites from patients with cirrhosis" J.Biochem. 109. 14-18 (1991)
[Publications] Yumiko TOMITA: "Stimulatory effect of pyruvate on hormonal induction of lipogenic enzymes in primary cultured hepatocytes" Biochim.Biophys.Acta. (1991)
[Publications] Koichi MATSUDA: "Identification of glucocorticoidー and cyclic AMP responsive elements of the rat serine dehydratase gene:Difference in responses of the transfected and chromosomal genes" Biochem.Biophys.Res.Commun.(1991)
[Publications] Tsukasa IGAWA: "Hepatocyte growth factor is a potent mitogen for cultured rabbit renal tubular epithelial cells" Biochem.Biophys.Res.Commun.174. 831-838 (1991)
[Publications] Yumiko TOMITA: "Change of gene expression between spheroid and monolayer cultures of rat hepatocytes" Biochem.Biophys.Res.Commun.(1991)

1990 Fiscal Year Annual Research Report

培養肝細胞を用いた機能と増殖を調節する肝組織構築の生化学的研究

Principal Investigator

市原 明 徳島大学, 酵素科学研究センター, 教授 (40035374)

Research Products

[Publications] Akira ICHIHARA: "Regulation of protein and amino acid metabolisms in cultured hapatocytes" Nutrion:Protein and Amino Acids. 49-66 (1990)

[Publications] Chiseko NODA: "Organization and strucutre of 5'flanking region of the rat serine dehydratase gene" J.Biochem.108. 622-628 (1990)

[Publications] Chiseko NODA: "Developmental and growth related regulation of expression of serine dehydratase mRNA in liver" Biochem.Biophys.Res.Commun.168. 335-342 (1990)

[Publications] Keiji TANAKA: "Proteasomes (Multicatalytic proteinase complex) in eukaryotic cells" Cell Struct.Funct.15. 127-132 (1990)

[Publications] Chizuko Tsurumi: "cDNA cloning and sequencing for the import precursor of subunit B H^+ーATP synthase from rat mitochondria" Biochem.Biophys.Res.Commun.169. 136-142 (1990)

[Publications] Tomihiko HIGUCHI: "cDNA cloning and sequencing for the import precursor of coupling factor 6 in H^+ーATP synthase from rat liver mitochondria" Biochem.Biophys.Res.Commun.171. 1079-1086 (1990)

[Publications] Osamu ASAMI: "Purification and characterization of hepatocyte growth factor from injured liver of carbon tetrachloride treated rats" J.Biochem.109. 8-31 (1991)

[Publications] Shigeru YOSHIZAWA: "Inhibition of protein phosphatases by microcyctin and nodularin associated with hepatotoxicity" J.Cancer Clin Oncol.116. 609-614 (1990)

[Publications] Ichiro SHIMIZU: "Hepatocyte growth factor in ascites from patients with cirrhosis" J.Biochem. 109. 14-18 (1991)

[Publications] Yumiko TOMITA: "Stimulatory effect of pyruvate on hormonal induction of lipogenic enzymes in primary cultured hepatocytes" Biochim.Biophys.Acta. (1991)

[Publications] Koichi MATSUDA: "Identification of glucocorticoidー and cyclic AMP responsive elements of the rat serine dehydratase gene:Difference in responses of the transfected and chromosomal genes" Biochem.Biophys.Res.Commun.(1991)

[Publications] Tsukasa IGAWA: "Hepatocyte growth factor is a potent mitogen for cultured rabbit renal tubular epithelial cells" Biochem.Biophys.Res.Commun.174. 831-838 (1991)

[Publications] Yumiko TOMITA: "Change of gene expression between spheroid and monolayer cultures of rat hepatocytes" Biochem.Biophys.Res.Commun.(1991)

市原明徳島大学, 酵素科学研究センター, 教授 (40035374)