Study of newly autologous chondrocyte implantation using by gene transfer of hyaluronan synthase

透明质酸合酶基因转移新自体软骨细胞移植的研究

• 批准号: 13671491
• 负责人: WADA Yuichi
• 金额: $ 2.18万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13671491/
• 关键词: hyaluronan; gene transfer; chondrocyte implantation; ATDC5; retrovirus vector; ヒアルロン酸合成酵素

In the present study, plasmids containg HAS2 under the PMX-IRES-GFP retrovirus vector were transfected into undifferentiated ATDC5. As a control, cells transfected with the empty vector were used. ATDC5 cells differentiated in the presence of insulin and RT-PCR was performed to detect HAS2 m RNA expression and we evaluated visualization of pericellular coats which were composed of newly synthesized HA using by immunofluorescent staining and analyzed HA concentration in culture medium by high performance liquid chromatography. The signal of the cells transfected with HAS2 was visually much stronger than that of control cells. The HA concentration of the cells transfected with HAS2 in culture medium was much higher than that of control throughout 21 days. By transfection of HAS2 with retrovirus, we are able to expect the continuous synthesis of HA. Therefore this technique may be a great significance in autologous chondrocyte implantation.

在本研究中，将含有HAS 2的质粒在PMX-IRES-GFP逆转录病毒载体下转染未分化的ATDC 5。作为对照，使用用空载体转染的细胞。用RT-PCR检测胰岛素诱导分化的ATDC 5细胞HAS 2 mRNA的表达，免疫荧光染色观察细胞外膜的变化，高效液相色谱分析培养液中HA的浓度。用HAS 2转染的细胞的信号在视觉上比对照细胞的信号强得多。转染HAS 2的细胞在培养液中的HA浓度在21天内明显高于对照组。通过用逆转录病毒转染HAS 2，我们能够预期HA的连续合成。因此，该技术在自体软骨细胞移植中具有重要意义。

项目成果

K.Tsuchiya, Y.Wada, K.Takase, K.Nakagawa, T.Yamashita, A.Watanabe and H.Moriya: "EVALUATION OF HYALURONAN SYNTHESIS AND EXPRESSION OF CD44 BY GENE TRANSFER OF HYALURONAN SYNTHASE 2"Osteoarthritis and Cartilage. 10 supplement B. (2002)

K.Tsuchiya、Y.Wada、K.Takase、K.Nakakawa、T.Yamashita、A.Watanabe 和 H.Moriya：“通过透明质酸合成酶 2 的基因转移评估透明质酸合成和 CD44 的表达”骨关节炎和软骨。

Kan Tsuchiya ,Yuichi Wada, Kan Takase, Kouichi Nakagawa, Takeshi Yamashita and Hideshige Moriya: "EVALUATION OF HYALURONAN SYNTHESIS BY RETROVIRUS-MEDIATED GENE TRANSFER OF HYALURONAN 2"J. Jpn. Orthop. Assoc.. 76(8). S1095 (2002)

Kan Tsuchiya、Yuichi Wada、Kan Takase、Kouichi Nakakawa、Takeshi Yamashita 和 Hideshige Moriya：“通过逆转录病毒介导的透明质酸 2 基因转移来评估透明质酸合成”J。

Kan Tsuchiya, Yuichi Wada, et al.: "EVALUATION of HYALURONAN SYNTHESIS BY RETROVIRUS-MEDIATED GENE TRANSFER of HYALURONAN"Osteoarthritis and Cartilage. Vol.10. SB (2002)

Kan Tsuchiya、Yuichi Wada 等人：“通过逆转录病毒介导的透明质酸基因转移来评估透明质酸合成”骨关节炎和软骨。

土屋敢, 和田佑一, 高瀬完, 他: "レトロウイルスを用いたヒアルロン酸合成酵素遺伝子導入によるヒアルロン酸合成量の評価"日本整形外科学会雑誌. Vol.76 No.8. S1095 (2002)

Tsuyoshi Tsuchiya、Yuichi Wada、Kan Takase 等：“利用逆转录病毒导入透明质酸合酶基因来评价透明质酸合成量”日本骨科学会杂志 Vol.76 No.8 (2002)。

Kan Tsuchiya, Yuichi Wada, et al.: "Evaluation of Hyaluronan Synthesis by Retrovirus-Mediated Gene Transfer of Hyaluronan"Osteoarthritis and Cartilage. Vol.10. S.B. (2002)

Kan Tsuchiya、Yuichi Wada 等人：“通过逆转录病毒介导的透明质酸基因转移来评估透明质酸合成”骨关节炎和软骨。