title: Role of nitrite, urate and pepsin in the gastroprotective effects of saliva.
abstract: Dietary nitrate is now recognized as an alternative substrate for nitric oxide (•NO) production in the gut. This novel pathway implies the sequential reduction of nitrate to nitrite, •NO and other bioactive nitrogen oxides but the physiological relevance of these oxidants has remained elusive. We have previously shown that dietary nitrite fuels an hitherto unrecognized nitrating pathway at acidic gastric pH, through which pepsinogen is nitrated in the gastric mucosa, yielding a less active form of pepsin in vitro. Here, we demonstrate that pepsin is nitrated in vivo and explore the functional impact of protein nitration by means of peptic ulcer development. Upon administration of pentagastrin and human nitrite-rich saliva or sodium nitrite to rats, nitrated pepsin was detected in the animal's stomach by immunoprecipitation. •NO was measured in the gastric headspace before and after nitrite instillation by chemiluminescence. At the end of each procedure, the stomach's lesions, ranging from gastric erosions to haemorrhagic ulcers, were scored. Nitrite increased gastric •NO by 200-fold (p<0.05) and nitrated pepsin was detected both in the gastric juice and the mucosa (p<0.05). Exogenous urate, a scavenger of nitrogen dioxide radical, blunted •NO detection and inhibited pepsin nitration, suggesting an underlining free radical-dependent mechanism for nitration. Functionally, pepsin nitration prevented the development of gastric ulcers, as the lesions were only apparent when pepsin nitration was inhibited by urate. In sum, this work unravels a novel dietary-dependent nitrating pathway in which pepsin is nitrated and inactivated in the stomach, preventing the progression of gastric ulcers.
authors.affiliation: Faculty of Pharmacy and Center for Neurosciences and Cell Biology, University of Coimbra, Health Sciences Campus, Azinhaga de Santa Comba, 3000-548 Coimbra, Portugal. Electronic address: barbarasrocha@icloud.com.
authors.forename: Bárbara S
authors.initials: BS
authors.lastname: Rocha
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institute, Stockholm, Sweden.
authors.forename: Jon O
authors.initials: JO
authors.lastname: Lundberg
authors.affiliation: Departamento de Bioquímica, Facultad de Medicina, Universidad de La República, Montevideo, Uruguay; Center for Free Radical and Biomedical Research, Facultad de Medicina, Universidad de La República, Montevideo, Uruguay.
authors.forename: Rafael
authors.initials: R
authors.lastname: Radi
authors.affiliation: Faculty of Pharmacy and Center for Neurosciences and Cell Biology, University of Coimbra, Health Sciences Campus, Azinhaga de Santa Comba, 3000-548 Coimbra, Portugal.
authors.forename: João
authors.initials: J
authors.lastname: Laranjinha
mesh_terms: D000818:Animals; D005609:Free Radicals / Q000378:metabolism; D005753:Gastric Mucosa / Q000378:metabolism; D006801:Humans; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism*; D016877:Oxidants / Q000378:metabolism; D010434:Pepsin A / Q000378:metabolism*; D020116:Pepsinogen A / Q000378:metabolism; D051381:Rats; D012463:Saliva / Q000378:metabolism*; D014527:Uric Acid / Q000378:metabolism*
chemical_list: D005609:Free Radicals; D009566:Nitrates; D009573:Nitrites; D016877:Oxidants; D014527:Uric Acid; D009569:Nitric Oxide; D020116:Pepsinogen A; D010434:Pepsin A
keywords: Nitrate; Nitrite; Stomach; Tyrosine nitration