| Research Abstract |
The dystrophic form of epidermolysis bullosa (DEB) is characterized by subliminal densa blister, and mutations in the type VII collagen gene have been shown to underlie DEB. To address gene therapy of DEB, we tried to introduce type VII collagen gene to keratinocytes. We first cloned a 9-kb full-length cDNA of the type VII collagen gene, constructed a expression vector of the cDNA and introduced it to rat kerationcytes using the naked DNA method. The results demonstrated that the expression of transgene was found in kerationcytes 24 h later and in basement membrane zone a week later. Success in expression of this collagen suggested a possibility of gene therapy for DEB.
Also, we constructed an IL-6 expression vector and introduced it to rat keratinocytes. Introduction of the IL-6 gene induced erythema macroscopically, and epidermal hypertrophy and lymphocytic infiltration microscopically. Next, we could show that pre-introduction of IL-6 antagonsit genes to keratinocytes inhibited inflammation by post-introduction of the IL-6 gene. Since IL-6 production was increased in keratinocytes of psoriasis, our result indicated possibility of gene therapy for psoriasis. In keratinocytes of psoriasis. Furthermore, we introduced IL-10 gene to keratinocytes and then measured serum level of IL-10. Significant level of IL-10 was found in serum and circulating IL-10 inhibited contact hypersensitivity at distant areas of the skin. This result suggested gene therapy for systemic diseases using keratinocyte as a bioreactor.
Besides the above results, we develop new gene transfer methods to keratinocytes and found several mutations in patients with DEB.
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