Background: Interleukin-13 (IL-13) has been implicated in the pathogenesis of fibrotic conditions. Previously, a murine model for scleroderma has been established by repeated local injections of bleomycin. This animal model enabled us to study local expression and production of IL-13 in skin lesions during disease progression. Methods: Dermal sclerosis (DSc) was induced by repeated subcutaneous injections of bleomycin (1 mg/ml) in C3H/HeJ mice. IL-13 and IL-4 expressions were examined by RT-PCR, ELISA and immunohistochemistry. Results: RT-PCR showed that both IL-4 and IL-13 mRNA levels in skin lesions were increased and peaked after 4 weeks of bleomycin treatment. Quantification by densitometry revealed up to 4.2- and 1.9-fold increases, respectively. Immunohistochemical localization showed in skin lesions expression of IL-13 on infiltrating inflammatory cells, including mononuclear cells and possibly mast cells, increased with DSc progression. IL-13 protein production was also significantly increased. In skin lesions, IL-13 receptor (IL-13R) α2 expression was augmented mainly in the infiltrating mononuclear cells after 4 weeks of bleomycin exposure. IL-13Rα2, but not IL-13Rα1, mRNA was upregulated in the whole skin after 4 weeks. On the contrary, mRNA expression of IL-13Rα1 and IL- 13Rα2 was significantly altered in the cultured fibroblasts derived from bleomycin-treated skin. Conclusion: These data demonstrate that in skin lesions levels of IL-13 as well as its receptor increase in parallel with DSc progression, suggesting that IL-13 promotes the progression of cutaneous fibrosis/sclerosis in the murine model of bleomycin-induced scleroderma.
SummaryAccumulative data have demonstrated that plasminogen activator inhibitor-1 (PAI-1) plays an important role in the extracellular matrix metabolism; however, the involvement of PAI-1 in scleroderma has not been fully elucidated. In this study, we investigated the role of PAI-1 in bleomycin-induced murine scleroderma. 100 m m m m g of bleomycin was injected subcutaneously to the back skin of C3H/HeJ mice on alternate day for 4 weeks. Histopathological findings revealed that PAI-1 was positive in macrophage-like cells and fibroblastic cells in the dermis, in parallel with the induction of dermal sclerosis. PAI-1 mRNA expression in the whole skin was up-regulated at 1 and 4 weeks. The production of active PAI-1 protein in the lesional skin was significantly increased 3 and 4 weeks after bleomycin treatment. Next, we examined whether dermal sclerosis is induced by bleomycin in PAI-1-deficient (PAI-1-/-) mice. 10 m m m m g of bleomycin was subcutaneously injected to PAI-1-/-and wild type (WT) mice 5 days per week for 4 weeks. Histological examination revealed that dermal sclerosis was similarly induced even in PAI-1-/-as well as WT mice. Dermal thickness and collagen contents in the skin were significantly increased by bleomycin injection in both PAI-1-/-and WT mice, and the rate of increase was similar. These data suggest that PAI-1 plays an important role, possibly via TGF-b b b b pathway activation. However, the fact that PAI-1 deficiency did not ameliorate skin sclerosis suggest that PAI-1 is not the essential factor in the development of bleomycin-induced scleroderma, and more complex biochemical effects other than PA/plasmin system are greatly suspected.
A method for the quantitative evaluation of topically applied anti-inflammatory agents is described. Conjunctival inflammation was induced in rabbits by topical instillation of n-butanol. The intensity of inflammation was determined by measuring changes of corneal surface temperature with an infrared thermometer. The closest correlation was obtained between corneal temperature change and the Draize score which is widely used as a subjective scoring method. Dexamethasone showed a good logarithmic dose-response inhibitory effect between 0.0001 and 0.1 %, and glycyrrhizin the same at 0.25–5%. Glycyrrhizin in a 5% solution showed a comparable anti-inflammatory effect to that of dexamethasone (0.1%). The inflammation model induced by n-butanol was mediated, in part, by the degranulation of mast cells because of some inhibitory effect of disodium cromoglycate (2%), an inhibitor of mast cell degranulation, and diphenhydramine hydrochloride (0.5%), an antihistaminic agent.
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