Nitric oxide synthase isoforms expression in fibroblasts isolated from human normal peritoneum and adhesion tissues

ObjectiveTo determine the expression of nitric oxide synthases (NOSs) and their modulation by hypoxia in human peritoneal (NF) and adhesion fibroblasts (ADF).DesignProspective experimental study.SettingUniversity medical center.Patient(s)Fibroblasts from peritoneum and adhesion tissues.Intervention(s)Hypoxia and silencing inducible NOS (iNOS) gene expression in fibroblasts.Main Outcome Measure(s)We used reverse-transcriptase polymerase chain reaction to quantify messenger RNA (mRNA) levels of NOS isoforms. Griess assay was used to measure NO levels.Result(s)The mRNA copies/μg RNA of neuronal NOS (nNOS) and endothelial NOS (eNOS) were 6.6 × 103 in NF, 5.7 × 103 in ADF and 7.0 × 103 in NF, 6.1 × 103 in ADF, respectively. The mRNA copies/μg RNA of iNOS were 31.3 × 103 in NF and 33.0 × 103 in ADF. Hypoxia increased iNOS mRNA copies/μg RNA from 31.3 × 103 to 61.3 × 103 in NF and from 33.0 × 103 to 63.9 × 103 in ADF, whereas there were no changes in mRNA levels of nNOS and eNOS in NF and ADF. Nitric oxide levels were lower in ADF (0.94 μmol/L) than NF (1.97 μmol/L). Silencing iNOS decreased NO levels in NF (from 1.97 μmol/L to 0.41 μmol/L) and in ADF (from 0.94 μmol/L to 0.27 μmol/L).Conclusion(s)Nitric oxide synthases are differentially expressed in NF and ADF, with iNOS being the most expressed and the main source of NO. Hypoxia was shown to alter the expression of NOSs and NO in NF and ADF.