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Reactive oxygen and nitrogen metabolites modulate fibronectin-induced fibroblast migration in vitro
1Research Service, Southern Arizona Veterans Health Care System, Tucson, AZ 85723, USA.
Abstract:
Nitration of proteins by peroxynitrite may alter protein function. We hypothesized that reactive nitrogen species modulate fibronectin-induced fibroblast migration. To test this hypothesis, we evaluated fibroblast migration induced by fibronectin incubated with and without peroxynitrite. Peroxynitrite attenuated fibronectin-induced fibroblast migration in a dose-dependent manner but did not attenuate complement-activated serum-induced fibroblast migration. The reducing agents, deferoxamine and dithiothreitol (DTT), and L-tyrosine reversed the inhibition by peroxynitrite. PAPA-NONOate, a nitric oxide (NO) donor, and superoxide generated by the action of xanthine oxidase on lumazine or xanthine, also showed an inhibitory effect on fibroblast migration. The peroxynitrite generator, 3-morpholinosydnonimine (SIN-1), caused a concentration-dependent inhibition of fibroblast migration. Peroxynitrite reduced fibronectin binding to fibroblasts and resulted in nitrotyrosine formation. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of fibronectin binding to fibroblasts and suggest that peroxynitrite may play a role in regulation of fibroblast migration.
Insights
Reactive nitrogen species, like peroxynitrite, inhibit fibroblast migration by reducing fibronectin binding. This suggests peroxynitrite regulates fibroblast movement and may impact wound healing.
Area of Science:
- Biochemistry
- Cell Biology
- Immunology
Background:
- Protein nitration by peroxynitrite can alter protein function.
- Reactive nitrogen species (RNS) are implicated in various biological processes.
- Fibroblast migration is crucial for tissue repair and wound healing.
Purpose of the Study:
- To investigate the hypothesis that RNS modulate fibronectin-induced fibroblast migration.
- To determine the effect of peroxynitrite on fibroblast migration.
- To elucidate the mechanism by which peroxynitrite affects fibronectin-mediated cell movement.
Main Methods:
- Fibroblast migration assays were performed using fibronectin incubated with and without peroxynitrite.
- The effects of various agents (reducing agents, NO donors, superoxide generators) on migration were evaluated.
- Fibronectin binding to fibroblasts and nitrotyrosine formation were assessed.
Main Results:
- Peroxynitrite dose-dependently attenuated fibronectin-induced fibroblast migration.
- This inhibition was specific to fibronectin-induced migration, not serum-induced migration.
- Peroxynitrite reduced fibronectin binding to fibroblasts and led to nitrotyrosine formation, which was reversed by reducing agents and L-tyrosine.
Conclusions:
- Peroxynitrite inhibits fibronectin-induced fibroblast migration, likely through nitration of tyrosine residues.
- This mechanism impairs fibronectin binding to fibroblasts.
- Peroxynitrite may play a regulatory role in fibroblast migration, impacting tissue repair processes.