2003 Fiscal Year Annual Research Report

小型肝細胞の特異的表面抗原の同定と体外環境における肝組織ユニット構築

Research Project

Project/Area Number	14370393
Research Institution	Sapporo Medical University
Principal Investigator	三高俊広札幌医科大学, 附属がん研究所, 教授 (50231618)
Co-Investigator(Kenkyū-buntansha)	平田公一札幌医科大学, 医学部, 教授 (50136959) 吉川大和札幌医科大学, 附属がん研究所, 助手 (20274227) 竹田寛札幌医科大学, 附属がん研究所, 助手 (00333310) 水口徹札幌医科大学, 医学部, 助手 (30347174) 桂巻正札幌医科大学, 医学部, 助教授 (50253993)
Keywords	小型肝細胞 / 幹細胞 / 凍結保存 / ヒト肝細胞 / Scaffold / 肝組織構築 / 人工肝臓 / 薬物代謝酵素
Research Abstract	我々は、分離した小型肝細胞コロニーをコラーゲンスポンジ上に播種することにより、迅速に類肝組織構造を誘導することに成功した。約2週間で成熟肝細胞と胆管類似構造を持つ類肝組織を形成することができた。この組織は、肝細胞の分化機能であるアルブミン分泌能や尿素合成能、アンモニア代謝能を持っており、約一ヶ月間その機能を維持した。また、外科的に切除されたヒト正常肝組織から分離した肝細胞を長期間培養することができた。この系で培養するとラット小型肝細胞によく似た細胞が増殖し、一部は細胞が盛り上がり3次元的な構築をするようになった。さらにヒト肝臓から分離した細胞を直接コラーゲンスポンジ上に播種することにより、胆管上皮細胞と成熟肝細胞を有する組織を形成させることに成功した。胆管上皮細胞は、スポンジの表層を覆うように一層に配列するものとスポンジ内で管腔構造を構築するものとがみられた。表層の胆管上皮細胞の一部は、肝細胞マーカーを発現しており、肝細胞由来の可能性があった。培液中にはアルブミンがよく分泌されていた。半年以上凍結保存したラット小型肝細胞を培養し、成熟化させると薬剤に対応したCYP蛋白質を誘導できることが分かった。誘導されたCYPは酵素活性も有していた。ヒト小型肝細胞を同様に分離培養することにより、薬物代謝機能をin vitroで測定するなどの応用が期待される。ラット小型肝細胞を多孔性膜上で培養し、コロニーを作らせた後、細胞同士が重なるように2枚の膜を重ねると、細胞同士が接着し、成熟化させることができた。成熟肝細胞同士では不可能なので小型肝細胞に特異的な現象と考えられる。積層させることにより小型肝細胞の成熟化が促進され、細胞間に毛細胆管がよく形成されるようになる。類肝組織を組み込んだ人工肝臓のモデルになると考えている。今後は、ヒト小型肝細胞を組み込んだモデルを作りたいと考えている。

Research Products

(21 results)

All Other

All Publications (21 results)

[Publications] Shinichiro Ikeda, et al.: "Tumor necrosis factor-α and IL-6 reduce bile canalicular contractions of rat hepatocytes"Surgery. 133(1). 101-109 (2003)
[Publications] Atsushi Takamura, et al.: "Accumulation of Hsp70/Hsc70 molecular chaperone regulator BAG-1 on COPI-coated structures in gastric epithelial cells"International Journal of Oncology. 23(5). 1301-1308 (2003)
[Publications] Keisuke Harada, et al.: "Rapid formation of hepatic organoid in collagen sponge by rat small hepatocytes and hepatic nonparenchymal cells"Journal of Hepatology. 39(11). 716-723 (2003)
[Publications] 三高俊広, 他: "In Vitroにおける肝組織形成"最新医学. 58(3). 708-718 (2003)
[Publications] 杉本真一, 他: "肝幹細胞研究の現状〜小型肝細胞〜"肝胆膵. 46(3). 327-333 (2003)
[Publications] Yin Y, et al.: "Expression of laminin chains by central neurons : analysis with gene and protein trapping techniques"Genesis. 36(2). 114-127 (2003)
[Publications] Ferletta M, et al.: "Opposing roles of integrin alpha6betal and dystroglycan in laminin-mediated extracellular signal-regulated kinase activation"Molecular and Biological Cell. 14(5). 2088-2103 (2003)
[Publications] Yamato Kikkawa, et al.: "Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the glomerular basement membrane"Journal of Cell Biology. 161(1). 187-196 (2003)
[Publications] Yasutoshi Kimura, et al.: "Frequent beta-catenin alteration in gallbladder carcinomas"Journal of Experimental Clinical Cancer Research. 22(2). 321-328 (2003)
[Publications] H.Tanaka, et al.: "Differential gene expression screening between parental and highly metastatic pancreatic cancer variants using a DNA microarray"Journal of Experimental Clinical Cancer Research. 22(2). 307-313 (2003)
[Publications] Masayo Kawakami, et al.: "Expression analysis of vascular endothelial growth factors and their relationships to lymph node metastasis in human colorectal cancer"Journal of Experimental Clinical Cancer Research. 22(2). 229-237 (2003)
[Publications] Hitoshi Kimura, et al.: "Role of inducible nitric oxide synthase in pig liver transplantation"Journal of Surgical Research. 111(1). 28-37 (2003)
[Publications] Masayo Kawakami, et al.: "Quantification of vascular endothelial growth factor-C and its receptor-3 messenger RNA with real-time quantitative polymerase chain reaction as a predictor of lymph node metastasis in human colorectal cance"Surgery. 133(3). 300-308 (2003)
[Publications] Hiroomi Tatsuumi, et al.: "Surgical influence on TH1/TH2 balance and monocyte surface antigen expression and its relation to infectious complications"World Journal of Surgery. 27(5). 522-528 (2003)
[Publications] Makoto Meguro, et al.: "Apoptosis and necrosis after warm ischemia-reperfusion injury of the pig liver and their inhibition by ONO-1714"Transplantation. 75(5). 703-710 (2003)
[Publications] Yasutoshi Kimura, et al.: "Traumatic transection of the pancreas head : report of a case with santorini duct injury treated by damage-control surgery"Pancreas. 26(2). 205-207 (2003)
[Publications] Akihiro Nui, et al.: "Successful ex vivo normothermic liver perfusion with purely artificial products using artificial blood"International Journal of Artificial Organ. 26(1). 46-52 (2003)
[Publications] Tomohisa.Furuhata, et al.: "Efficacy of SPIO-MR imaging in the diagnosis of liver metastases from colorectal carcinomas"Diagnostic Surgery. 20(4). 321-325 (2003)
[Publications] Tadashi Katsuramaki, et al.: "Preoperative estimation of risk in hepatectomy using technetium-99m-galactosyl human serum albumin receptor amount by nonlinear 3-compartment model"Hepatogastroenterology. 50(49). 174-177 (2003)
[Publications] 原田啓介, 他: "肝臓の再生医学"顕微鏡. (印刷中). (2004)
[Publications] Ryo Sudo, et al.: "Bile canalicular formation in hepatic organoid reconstructed by rat small hepatocytes and hepatic nonparenchymal cells"Journal of Cellular Physiology. (in press). (2004)

2003 Fiscal Year Annual Research Report

小型肝細胞の特異的表面抗原の同定と体外環境における肝組織ユニット構築

Principal Investigator

三高 俊広 札幌医科大学, 附属がん研究所, 教授 (50231618)

Research Products

[Publications] Shinichiro Ikeda, et al.: "Tumor necrosis factor-α and IL-6 reduce bile canalicular contractions of rat hepatocytes"Surgery. 133(1). 101-109 (2003)

[Publications] Atsushi Takamura, et al.: "Accumulation of Hsp70/Hsc70 molecular chaperone regulator BAG-1 on COPI-coated structures in gastric epithelial cells"International Journal of Oncology. 23(5). 1301-1308 (2003)

[Publications] Keisuke Harada, et al.: "Rapid formation of hepatic organoid in collagen sponge by rat small hepatocytes and hepatic nonparenchymal cells"Journal of Hepatology. 39(11). 716-723 (2003)

[Publications] 三高 俊広, 他: "In Vitroにおける肝組織形成"最新医学. 58(3). 708-718 (2003)

[Publications] 杉本 真一, 他: "肝幹細胞研究の現状〜小型肝細胞〜"肝胆膵. 46(3). 327-333 (2003)

[Publications] Yin Y, et al.: "Expression of laminin chains by central neurons : analysis with gene and protein trapping techniques"Genesis. 36(2). 114-127 (2003)

[Publications] Ferletta M, et al.: "Opposing roles of integrin alpha6betal and dystroglycan in laminin-mediated extracellular signal-regulated kinase activation"Molecular and Biological Cell. 14(5). 2088-2103 (2003)

[Publications] Yamato Kikkawa, et al.: "Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the glomerular basement membrane"Journal of Cell Biology. 161(1). 187-196 (2003)

[Publications] Yasutoshi Kimura, et al.: "Frequent beta-catenin alteration in gallbladder carcinomas"Journal of Experimental Clinical Cancer Research. 22(2). 321-328 (2003)

[Publications] H.Tanaka, et al.: "Differential gene expression screening between parental and highly metastatic pancreatic cancer variants using a DNA microarray"Journal of Experimental Clinical Cancer Research. 22(2). 307-313 (2003)

[Publications] Masayo Kawakami, et al.: "Expression analysis of vascular endothelial growth factors and their relationships to lymph node metastasis in human colorectal cancer"Journal of Experimental Clinical Cancer Research. 22(2). 229-237 (2003)

[Publications] Hitoshi Kimura, et al.: "Role of inducible nitric oxide synthase in pig liver transplantation"Journal of Surgical Research. 111(1). 28-37 (2003)

[Publications] Masayo Kawakami, et al.: "Quantification of vascular endothelial growth factor-C and its receptor-3 messenger RNA with real-time quantitative polymerase chain reaction as a predictor of lymph node metastasis in human colorectal cance"Surgery. 133(3). 300-308 (2003)

[Publications] Hiroomi Tatsuumi, et al.: "Surgical influence on TH1/TH2 balance and monocyte surface antigen expression and its relation to infectious complications"World Journal of Surgery. 27(5). 522-528 (2003)

[Publications] Makoto Meguro, et al.: "Apoptosis and necrosis after warm ischemia-reperfusion injury of the pig liver and their inhibition by ONO-1714"Transplantation. 75(5). 703-710 (2003)

[Publications] Yasutoshi Kimura, et al.: "Traumatic transection of the pancreas head : report of a case with santorini duct injury treated by damage-control surgery"Pancreas. 26(2). 205-207 (2003)

[Publications] Akihiro Nui, et al.: "Successful ex vivo normothermic liver perfusion with purely artificial products using artificial blood"International Journal of Artificial Organ. 26(1). 46-52 (2003)

[Publications] Tomohisa.Furuhata, et al.: "Efficacy of SPIO-MR imaging in the diagnosis of liver metastases from colorectal carcinomas"Diagnostic Surgery. 20(4). 321-325 (2003)

[Publications] Tadashi Katsuramaki, et al.: "Preoperative estimation of risk in hepatectomy using technetium-99m-galactosyl human serum albumin receptor amount by nonlinear 3-compartment model"Hepatogastroenterology. 50(49). 174-177 (2003)

[Publications] 原田 啓介, 他: "肝臓の再生医学"顕微鏡. (印刷中). (2004)

[Publications] Ryo Sudo, et al.: "Bile canalicular formation in hepatic organoid reconstructed by rat small hepatocytes and hepatic nonparenchymal cells"Journal of Cellular Physiology. (in press). (2004)

三高俊広札幌医科大学, 附属がん研究所, 教授 (50231618)

[Publications] 三高俊広, 他: "In Vitroにおける肝組織形成"最新医学. 58(3). 708-718 (2003)

[Publications] 杉本真一, 他: "肝幹細胞研究の現状〜小型肝細胞〜"肝胆膵. 46(3). 327-333 (2003)

[Publications] 原田啓介, 他: "肝臓の再生医学"顕微鏡. (印刷中). (2004)