Results, meta-analysis and a first evaluation of UNOxR, the urinary nitrate-to-nitrite molar ratio, as a measure of nitrite reabsorption in experimental and clinical settings

Abstract

We recently found that renal carbonic anhydrase (CA) is involved in the reabsorption of inorganic nitrite (NO), an abundant reservoir of nitric oxide (NO) in tissues and cells. Impaired NO synthesis in the endothelium and decreased NO bioavailability in the circulation are considered major contributors to the development and progression of renal and cardiovascular diseases in different conditions including diabetes. Isolated human and bovine erythrocytic CAII and CAIV can convert nitrite to nitrous acid (HONO) and its anhydride NO which, in the presence of thiols (RSH), are further converted to S-nitrosothiols (RSNO) and NO. Thus, CA may be responsible both for the homeostasis of nitrite and for its bioactivation to RSNO/NO. We hypothesized that enhanced excretion of nitrite in the urine may contribute to NO-related dysfunctions in the renal and cardiovascular systems, and proposed the urinary nitrate-to-nitrite molar ratio, i.e., UR, as a measure of renal CA-dependent excretion of nitrite. Based on results from clinical and experimental animal studies, here, we report on a first evaluation of UR. We determined UR values in preterm neonates, healthy children, and adults, in children suffering from type 1 diabetes mellitus (T1DM) or Duchenne muscular dystrophy (DMD), in elderly subjects suffering from chronic rheumatic diseases, type 2 diabetes mellitus (T2DM), coronary artery disease (CAD), or peripheral arterial occlusive disease (PAOD). We also determined UR values in healthy young men who ingested isosorbide dinitrate (ISDN), pentaerythrityl tetranitrate (PETN), or inorganic nitrate. In addition, we tested the utility of UR in two animal models, i.e., the LEW.1AR1-iddm rat, an animal model of human T1DM, and the APOE*3-Leiden.CETP mice, a model of human dyslipidemia. Mean UR values were lower in adult patients with rheumatic diseases (187) and in T2DM patients of the DALI study (74) as compared to healthy elderly adults (660) and healthy young men (1500). The intra- and inter-variabilities of UR were of the order of 50% in young and elderly healthy subjects. UR values were lower in black compared to white boys (314 vs. 483, P = 0.007), which is in line with reported lower NO bioavailability in black ethnicity. Mean UR values were lower in DMD (424) compared to healthy (730) children, but they were higher in T1DM children (1192). ISDN (3 × 30 mg) decreased stronger UR compared to PETN (3 × 80 mg) after 1 day (P = 0.046) and after 5 days (P = 0.0016) of oral administration of therapeutically equivalent doses. In healthy young men who ingested NaNO (0.1 mmol/kg/d), UR was higher than in those who ingested the same dose of NaCl (1709 vs. 369). In LEW.1AR1-iddm rats, mean UR values were lower than in healthy rats (198 vs. 308) and comparable to those in APOE*3-Leiden.CETP mice (151).

Activated L-Arginine/Nitric Oxide Pathway in Pediatric Cystic Fibrosis and Its Association with Pancreatic Insufficiency, Liver Involvement and Nourishment: An Overview and New Results

“…In our CF patients, we found higher (by 33%) U NOx R values than in the healthy controls, presumably due to a higher excretion rate of nitrate (by 28%) and a lower (by 17%), albeit insignificant, urinary nitrite excretion. The U NOx R values measured in the CF children are closely comparable with those measured in type I diabetic children [47]. CF children with insufficient pancreas function had higher (by 11%) U NOx R values than CF children with sufficient pancreas, while CF children with liver involvement had lower (by 13%) U NOx R values than CF children with insufficient pancreas function.…”

“…P NOx R and U NOx R correlated with each other in the CF patients, but they did not correlate with GFR. This observation may suggest that both the plasma and the urinary concentrations of nitrate and nitrite are primarily dependent upon the reabsorption of nitrite by carbonic anhydrase (CA) in the proximal tubule of the nephron [ 47 ] ( Figure 1 ). By using the drug acetazolamide, a strong inhibitor of CA activity, early work found that CA is involved in CF [ 60 , 61 , 62 ].…”

“…The urinary excretion of the NO metabolites nitrite and nitrate, of ADMA and its metabolite DMA was corrected for creatinine excretion and these data are reported as µM analyte per mM creatinine. Nitrate-to-nitrite (NOx) ratios (R) were calculated for plasma (P; P NOx R) and urine (U; U NOx R) samples [47].…”

Enhanced Nitric Oxide (NO) and Decreased ADMA Synthesis in Pediatric ADHD and Selective Potentiation of NO Synthesis by Methylphenidate

“…U NOx R is supposed to be a measure for the renal carbonic anhydrase-dependent reabsorption of nitrite. Carbonic anhydrase may also be responsible for NO formation in the presence of thiols such as L-cysteine and, thus, may determine the NO reservoir and bioavailability in the renal and cardiovascular systems [32]. Our study suggests that ritalinic acid, the major MPH metabolite, may have increased the reabsorption and/or decreased the excretion of urinary nitrite and nitrate by about 30% and 10%, respectively, thus eventually increasing the plasma nitrite concentration.…”

“…The nitrate-to-nitrite molar ratios in plasma (P NOx R) and urine (U NOx R) were determined by dividing the concentration of nitrate in plasma (P NO3 ) or urine (U NO3 ) by the concentration of nitrite in plasma (P NO2 ) or urine (U NO2 ) (P NOx R = P NO3 /P NO2 ; U NOx R = U NO3 /U NO2 ). U NOx R, P NOx R, and their ratio U NOx R/P NOx R are considered nitrite-dependent renal carbonic anhydrase activity [32]. The fractional excretion (FE, %) of nitrite and nitrate was calculated using the following formula: FE (%) = (U NOx × P Crea )/(P NOx × U Crea ) × 100.…”