Pharmaceutical Studies for Gene Therapy : Expression of Human Cu, Zn-Superoxide Dismutase Gene Transfected by Lipofection in Rat Skin Fibroblasts
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概要
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To evaluate whether lipofection using Lipofectin^<TM> is suitable for delivering foreign genes into skin fibroblasts as target cells, we performed experiments using human superoxide dismutase (hSOD) and neomycin-resistance (Neo^r) genes as models in rat skin fibroblasts (FR and primary cells) in vitro. The amounts of DNA used in the lipofection procedure significantly affected the transfection efficiencies, and the optimal amounts were determined for all cells used. However, the efficiencies in rat skin fibroblasts were about 20-fold higher than that in rat lung epithelial-like cells (L2 cells). The differences in plasmid vectors (pRc/RSV-SOD and pRc/CMV-SOD) hardly affected the transfection efficiencies. The amounts of Lipofectin^<TM> significantly affected the transfection efficiencies, and the optimal amounts were determined for both types of skin fibroblasts However, cytotoxic effects in both skin fibroblasts. were observed with high doses of Lipofectin^<TM>. On the other hand, with optimal amounts of DNA and Lipofectin^<TM>, the reporter gene (Neo^r) introduced into cells was mainly integrated into the host cell chromosome. Western blot analysis showed the continuous expression of hSOD protein for at least 45 d in skin fibroblasts transfected with the expression plasmid for hSOD by Lipofectin^<TM> under the optimal conditions, and the cellular SOD activity fluctuated in parallel with the expression of hSOD protein. Differences in the type of cells also affected the expression of hSOD. These results indicate that it is necessary to set up optimal conditions for transfection using Lipofectin^<TM> for each cell type, and that transfection with Lipofectin^<TM> under optimal conditions may be an efficient method for introduction of foreign genes into skin fibroblasts for use as a clinical delivery system of therapeutic protein.
- 公益社団法人日本薬学会の論文
- 1996-08-15
著者
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TANIGAWARA Yusuke
Department of Pharmacy, School of Medicine, Keio University
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OKUMURA Katsuhiko
Department of Hospital Pharmacy, Kobe University School of Medicine
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Nishiguchi Kohshi
Department of Hospital Pharmacy, School of Medicine, Kobe University
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IWAKAWA Seigo
Kobe Pharmaceutical University
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Maeda Tetsuo
Department of Internal Medicine, Harima Hospital of Ishikawajima-Harima Heavy Industries, Health Ins
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Okumura K
Department Of Clinical Evaluation Of Pharmacotherapy Kobe University Graduate School Of Medicine
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Maeda Tetsuo
Department Of Hematology And Oncology Graduate School Of Medicine Osaka University
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IWAKAWA Seigo
Department of Pharmaceutics, Kobe Pharmaceutical University
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KOMADA Fusao
Department of Drug Informatics, Faculty of Pharmaceutical Sciences, Josai University
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Kamiyama Fumio
京都大学再生医科学研究所
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Kodama Fusao
Cosmed Pharmaceutical Co. Ltd.
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Iwakawa S
Department Of Cardiology Miki
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Iwakawa Seigo
Department Of Hospital Pharmacy School Of Medicine Kobe University : Department Of Pharmacy Kyoto Un
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Tanigawara Yusuke
Department Of Clinical Pharmacokinetics And Pharmacodynamics School Of Medicine Keio University
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Nishiguchi K
Department Of Hospital Pharmacy Kobe University School Of Medicine
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Nishiguchi Kohshi
Department Of Clinical Pharmacy Faculty Of Pharmaceutical Sciences Kyoto Pharmaceutical University
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Nishiguchi Kohshi
Department Of Hospital Pharmacy School Of Medicine Kobe University
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ISHIDA Kazue
Kobe Pharmaceutical University
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NAKAJIMA Masanori
Department of Hospital Pharmacy, School of Medicine, Kobe University
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Okumura Katsuhiko
Department Of Clinical Evaluation Of Pharmacotherapy Kobe University Graduate School Of Medicine
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Okumura Katsuhiko
Department Of Hospital Pharmacy Kyoto Pharmaceutical University:department Of Hospital Pharmacy Kobe
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Ishida K
Kobe Pharmaceutical University
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Komada F
Department Of Pharmacy Kobe University School Of Medicine
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Shinobu Ichikawa
Department Of Cardiology Miki
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Nakamura Takeshi
Department Of Gastroenterological Surgery Kobe University Graduate School Of Medical Sciences
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Nagahiro Kazumi
Department Of Hospital Pharmacy Kobe University School Of Medicine
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Ichikawa Shinobu
Department Of Cardiology Miki City Hospital
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Maeda Tetsuo
Department Of Hospital Pharmacy School Of Medicine Kobe University
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Nakajima Masanori
Department Of Hospital Pharmacy School Of Medicine Kobe University
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Nakajima Masanori
Department Of Anthropology The University Of Tokyo
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Maeda Tetsuo
Department Of General Medicine Kanazawa University Hospital
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