Elements of the nitric oxide pathway can degrade TIMP-1 andincrease gelatinase activity

[摘要] Purpose: Keratoconus is a non-inflammatory thinning disorder of thecorneal stroma. Recently, we showed that these corneas contain induciblenitric oxide synthase and an accumulation of nitrotyrosine, representingoxidative damage from peroxynitrite. Previously, we suggested thatkeratoconus corneas and their cell cultures have alterations in agelatinase system with increased matrix metalloproteinase-2 (MMP-2)activity and decreased tissue inhibitor of metalloproteinase-1 (TIMP-1).This study examines whether a peroxynitrite donor (3-morpholinosydomineN-ethylcarbamide, SIN-1) or nitric oxide donor(S-nitroso-N-acetylpenicillamine, SNAP) could alter TIMP-1 and/or MMP-2in vitro.Methods: Normal stromal fibroblasts were cultured in the presence orabsence of either SIN-1 or SNAP for varying times. These cultures wereanalyzed by western and northern blot analyses, gelatin zymography, anda quantitative gelatinase/MMP assay.Results: In vitro, SIN-1 treatment led to protein nitration,increased RNA levels of TIMP-1 and MMP-2, and loss of TIMP-1immunostaining, but did not diminish gelatinase activity. SNAP treatmentled to activation of MMP-2 and significantly increased gelatinase/MMPactivity, without a change in TIMP-1 levels.Conclusions: Our data show that peroxynitrite or nitric oxide candecrease TIMP-1 and increase gelatinase activity, respectively. Thisdemonstrates a relationship between elements of oxidative stress andtissue degradation in human corneal fibroblasts. This effect may play asignificant role in the stromal thinning that occurs in keratoconus.

[学科分类] 生物化学/生物物理