Local and Systemic Alterations of the L-Arginine/Nitric Oxide Pathway in Sputum, Blood, and Urine of Pediatric Cystic Fibrosis Patients and Effects of Antibiotic Treatment

Abstract

Alterations in the L-arginine (Arg)/nitric oxide (NO) pathway have been reported in cystic fibrosis (CF; OMIM 219700) as the result of various factors including systemic and local inflammatory activity in the airways. The aim of the present study was to evaluate the Arg/NO metabolism in pediatric CF patients with special emphasis on lung impairment and antibiotic treatment. Seventy CF patients and 78 healthy controls were included in the study. CF patients (43% male, median age 11.8 years) showed moderately impaired lung functions (FEV1 90.5 ± 19.1% (mean ± SD); 21 (30%) had a chronic Pseudomonas aeruginosa (PSA) infection, and 24 (33%) had an acute exacerbation). Plasma, urinary, and sputum concentrations of the main Arg/NO metabolites, nitrate, nitrite, Arg, homoarginine (hArg), and asymmetric dimethylarginine (ADMA) were determined in pediatric CF patients and in healthy age-matched controls. Clinical parameters in CF patients included lung function and infection with PSA. Additionally, the Arg/NO pathway in sputum samples of five CF patients was analyzed before and after routine antibiotic therapy. CF patients with low fractionally exhaled NO (FENO) showed lower plasma Arg and nitrate concentrations. During acute exacerbation, sputum Arg and hArg levels were high and dropped after antibiotic treatment: Arg: pre-antibiotics: 4.14 nmol/25 mg sputum vs. post-antibiotics: 2.33 nmol/25 mg sputum, p = 0.008; hArg: pre-antibiotics: 0.042 nmol/25 mg sputum vs. post-antibiotics: 0.029 nmol/25 mg sputum, p = 0.035. The activated Arg/NO metabolism in stable CF patients may be a result of chronic inflammation. PSA infection did not play a major role regarding these differences. Exacerbation increased and antibiotic therapy decreased sputum Arg concentrations.

Keywords: Pseudomonas aeruginosa; antibiotics; cystic fibrosis; inflammation; nitric oxide.

Figure 1

Arg/NO metabolites in urine and plasma of healthy controls and CF patients grouped by FENO. (A–F) Arg/NO metabolites in plasma; (G–L) Arg/NO metabolites in urine. (A) Arginine (Arg) in plasma; (B,K) Asymmetric dimethylarginine (ADMA) in plasma and urine, respectively; (C) Arg/ADMA ratio in plasma; (D) Citrulline in plasma; (E,G) Nitrate in plasma and urine, respectively; (F,H) Nitrite in plasma and urine, respectively; (I) Nitrate/nitrite ratio in urine (J), Dimethylamine (DMA) in urine; (L) DMA/ADMA ratio in urine. Reduced FENO < 3rd percentile, normal FENO ≥ 3rd ≤ 95th percentile, increased FENO > 95th percentile. Data were available for 68 CF patients, FENO levels were missing in one patient, Arg/NO metabolites were not available for two patients in the blood and two patients in the urine. The number of subjects varied due to missing samples and small sample volumes in pediatric CF patients and controls. * p < 0.05; ** p < 0.001

Figure 2

Content (nmol analyte per 25 mg sputum) of the indicated Arg/NO metabolites in sputum samples of five CF patients before and after antibiotics intravenous infusion. (A) Arg, arginine; (B) ADMA, asymmetric dimethylarginine; (C) Arg/ADMA ratio, (D) nitrate; (E) nitrite; (F) S_NOxR nitrate/nitrite ratio in sputum, (G) hArg, homoarginine; (H) DMA, dimethylamine; (I) DMA/ADMA ratio.