2005 Fiscal Year Final Research Report Summary
Development of a new strategy for gene therapy of autosomal recessive congenital ichthyosis by gene transfer to the bulge stem cells
| Project/Area Number |
16390312
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Dermatology
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| Research Institution | Hokkaido University |
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Principal Investigator |
AKIYAMA Masashi Hokkaido Univ., Hokkaido University Hospital, Lecturer, 病院, 講師 (60222551)
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| Co-Investigator(Kenkyū-buntansha) |
SHIMIZU Hiroshi Hokkaido Univ., Graduate School of Medicine, Asso.Prof., 大学院・医学研究科, 教授 (00146672)
SAWAMURA Daisuke Hokkaido Univ., Graduate School of Medicine, Asso.Prof., 大学院・医学研究科, 助教授 (60196334)
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| Project Period (FY) |
2004 – 2005
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| Keywords | ichthyosis / hair follicle / gene therapy / mutation / bulge / stem cell / ichthyosis syndrome / transglutaminase |
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| Research Abstract |
Gene transfer to hair follicle, epithelium is an attractive approach for not only treating skin diseases, but also many systemic disorders. In this study, we intended to develop a gene transfer system for hair follicle epithelial stem cells to maximize the beneficial therapeutic effects. For persistent and stable transgene expression in hair follicle stem cells, we transferred retroviral vectors encoding reporter genes into cultured hair follicle stem cells. We performed gene transfection experiments into the bulge stem cells in vitro. We dissected bulge areas from mice vibrissa hair follicles and established primary cultures. We transfected the gene constructs that we had made last year. The transfected cells were mixed with cultured dermal papilla cells and transplanted on to immunodeficient mice. We succeeded in reconstituting hair follciles and their appendages in which these cells harbored a transgene reporter. The transgene expression was observed in all skin epithelial compartme
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nts including the hair follicle epithelium, sebaceous gland and epidermis. In addition, transgene expression was observed for at least 6 months.
In addition, we performed gene transfection experiments into the bulge stem cells in vivo using the methods that had been established from in vitro studies.
Furthermore, we performed transfection experiments of gene constructs for gene therapy. In detail, we cloned normal TGM1 cDNA construct and normal ABCA12 cDNA construct with the transfection vector and the reporter genes selected from the results of previous experiments. We obtained keratinocyte cultures form lamellar ichthyosis patients carrying TGM1 mutations and from harlequin ichthyosis patients harboring ABCA12 mutations after fully informed consents. Epidermis showing characteristic ichthyosis phenotype was reconstituted from these cultured keratinocytes form the patients on the back of nude mice. Targetting the reconstructed ichthyosis skin lesions, we tried gene therapy experiments using normal TGM1 or normal ABCA12 gene constructs made in the previous studies. This hair follicle stem cell targeted gene transfer and reconstitution system provides reliable gene-function analysis and gene therapy. Less
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