イヌの耳介軟骨から軟骨細胞を分離し、これを温度応答性培養皿上でシート状に増殖させ、シート状に回収した。本シートを積層化し、ラットよりコラゲナーゼ消化法にて分離した膵島を包埋し、三次元構造の人工膵を作製し、形態観察を行った。その結果、人工膵作製し、その安定化のために9日間培養後の組織像から、軟骨細胞中の膵島は活性を有しており、軟骨細胞シートを通じて栄養等が供給されていることが確認された。膵島は組織像上、完全に軟骨細胞層に封入されており、軟骨細胞シートが免疫隔離能を発揮する可能性が期待される。

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This report describes the primary culture of rat bone marrow cells. The hydroxyapatite (HA) microcarrier was used for the scaffold, which is compared with the HA disk and the T75 flask. Glucose consumption was measured to evaluate cell growth. Much higher glucose consumption was detected on the HA microcarrier group at the early stage of culture than on the T75 flask. This means that the cells on the HA microcarrier proliferated more rapidly. The differentiation potency to the osteoblast was evaluated to determine whether the proliferated cells on the HA microcarrier were still functionally active. The alkaline phosphatase activity and the osteocalcin production were detected, and the cells on the HA microcarrier maintained the differentiation potency to the osteoblasts. The three-dimensional cell culture using HA microcarrier is a useful procedure for clinical research in areas such as on artificial bone.

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HFDM-1 is a serum-free culture medium for fibroblasts. The amount of plasminogen activator inhibitor-1 (PAI-1) production from lung fibroblasts is closely related to the severity of the lung diseases. The growth rate of newly isolated human lung fibroblast and the amounts of PAI-1 of the cells were examined. Although the growth of fibroblasts cultured in HFDM-1, with or without 0.5% FCS, was slower than that in M199 containing 10% FCS during the first two or four days, respectively, the cells reached comparable numbers after six days. The expression of PAI-1 mRNA in the fibroblasts cultured with 0.5% FCS HFDM-1 was higher than in the cells cultured with 10% FCS M199. This effect was suppressed by the addition of gefitinib, the EGF signal blocker. There were no significant differences in the cellular PAI-1 expression among all three media. The PAI-1 activity in the culture medium and the fibroblasts’ surface uPA activity were suppressed when the cells were cultured in HFDM-1, with or without 0.5% serum, irrespective of the presence of gefitinib. These results suggested that HFDM-1 reduced the effects of serum on the fibroblasts’ PAI-1 production and activity, and would be a useful tool for the investigation of the lung fibroblasts’ roles in the pulmonary diseases.

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我々は Eagle’s Minimum Essential Medium（MEM）培地の製造会社が異なれば、ヒト肝細胞由来の不死化細胞（OUMS-２９）のコロニー増殖能が変わることを見出した。このことは、同じ組成のMEMを使用しても、製造会社の製品によって、実験結果が変わることを示唆している。培養細胞を使用して実験する研究者は注意すべきことであろう。

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