| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1996: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1995: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Research Abstract |
Nitric oxide (NO) is an intercellular messenger molecule involved in a variety of biological functions and is generated in mammalian organs by NO synthase (NOS). Inducidle NOS (iNOS) is induced in various types of tissues and cells by endotoxins, interferon-gamma (IFN-gamma), interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha), alone or in combination. In this study, we examined whether iNOS is induced in the epidermis of some skin diseases and in cytokine-stimulated keratinocytes. Isolated rat epidermal cells produced nitrite in the supernatant fluid when stimulated with interleukin-1beta and lipopolysaccharide. Depletion of Ia-positive cells from the epidermal cells slightly eliminated the activity of nitrite production. Homogenates of the stimulated cells exhibited a main band at 150kDa by the Western blot method. The same protein bands were also visible in the human keratinocyte cell line, HSC-1, cultured with the same stimulants. When stained with iNOS-specific monocl
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onal antibodies by the avidin-biotin-peroxidase immuno-histochemical method, the epidermis of some skin diseases, including lichenoid tissue reactions, psoriasis, collagen diseases and atopic dermatitis, showed a positive reaction to the antibodies. Topical medication of steroids decreased the expression with improvement of the disease activity in psoriasis and atopic dermatitis. There was no diffrence in iNOS expression in the epidermis between allergic type and croton-induced irritant type contact dermatitis. Peripheral blood cells had an ablity to stimulate keratinocytes to produce NO in atopic dermatitis patients compared to normal volunteers. These findings suggest that iNOS probably induced in keratincytes by cytokines is relevant to the pathology of some inflammatory skin diseases.
The nitrite production by UVB-irradiated kerationocytes and the effects of the NOS inhibitor on ear swelling and sunburn cell formation after UVB irradiation were examined. Nitrite was identified in the supernatant fluid of interleukin 1 (IL-1) -stimulated transformed keratinocyte cell lines. Five mJ/cm^2 UVB augumented the production of nitrite. Nitrite was not generated in the keratinocytes irradiated but not stimulated with IL-1. When BALB/c mice were irradiated with 200mJ/cm^2 UVB,ear swelling and sunburn cells were observed 24 hours after irradiation. When the mice were injected with a NOS inhibitor, N^G-nitro-L-arginine methyl ester (L-NAME), before irradiation, ear swelling was significantly suppressed.
Furthermore, such injection also suppressed the formation of sunburn cells. These findings suggest that NO produced in the skin by UVB irradiation contributes to keratinocyte injury as well as acute dermal reactions such as UVB edema. Less
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