NOS2 polymorphisms associated with the susceptibility to pulmonary arterial hypertension with systemic sclerosis: contribution to the transcriptional activity

Abstract

Systemic sclerosis (SSc) is a connective tissue disease characterized by tissue fibrosis. One of several complications of SSc, pulmonary arterial hypertension (PAH) can be refractory to treatment, both novel and established. In the present study we investigated the ratio of circulating nitric oxide to endothelin-1 in patients with both SSc and PAH, and determined whether polymorphisms in NOS2 (the nitric oxide synthase 2 gene) are associated with susceptibility to PAH. Endothelin-1 in plasma and nitric oxide metabolites (nitrate and nitrite) in serum were measured. The nitric oxide/endothelin-1 ratio was significantly lower in patients with both SSc and PAH than in patients with SSc only or in healthy control individuals. We confirmed the presence of two single nucleotide polymorphisms at positions -1,026 and -277 and a pentanucleotide repeat (CCTTT) at -2.5 kilobases. There were significant differences in single nucleotide polymorphisms between patients with SSc who had PAH and those who did not, and between patients with both SSc and PAH and healthy control individuals. The CCTTT repeat was significantly shorter in patients with both SSc and PAH than in patients with SSc only or in healthy control individuals. Transcriptional activity were analyzed using the luciferase reporter assay. The transcriptional activity of NOS2 was much greater in fibroblasts transfected by a vector with a long allele of the CCTTT repeat than in those transfected by a vector with a short allele. Polymorphisms in the NOS2 gene are associated with transcriptional activity of the NOS2 gene and with susceptibility to SSc-related PAH.

Unraveling the genetic component of systemic sclerosis

“…Recently, some loci have been proposed as PAH genetic markers, i.e., potassium voltage-gated channel shaker-related subfamily member 5, KCNA5 (associated with SSc and PAH) (WipV et al 2010); Endoglin, ENG (associated with PAH in SSc patients) (WipV et al 2007); Urokinase-type plasminogen activator receptor, uPAR (associated with PAH and also with digital ulcers appearance) (Manetti et al 2011); NOS2 (associated with PAH) (Kawaguchi et al 2006). …”

Pathogenic mechanisms of pulmonary arterial hypertension

“…Third, NO levels are intricately associated with oxidative stress, another likely but incompletely described regulator of pulmonary hypertension [135]. Finally, specific polymorphisms of NOS have been associated with pulmonary hypertension [136,137]; however, mechanistic details are lacking. Again, further characterization of the dysregulated action of NO and potential interactions with other pathogenic factors may depend upon the development of a genetically tractable animal model of severe PAH.…”

Genetics of systemic sclerosis-associated pulmonary arterial hypertension: Recent progress and current concepts

“…Although not tested in patients with connective tissue disease-associated PAH, these results are interesting, because ET-1 is a known modulator of Kv1.5 channels. Kawaguchi et al [46] found that the nitric oxide/ET-1 ratio was signifi cantly lower in patients with SSc and PAH compared with SSc patients without PAH and healthy controls. These investigators also found three SNPs in the NOS2 gene that were associated with SSc-associated PAH [46].…”