Sonic hedgehog protein regulates fibroblast growth factor 8 expression in metanephric explant culture from BALB/c mice: Possible mechanisms associated with renal morphogenesis.

The sonic hedgehog (SHH) morphogen regulates cell differentiation and controls a number of genes during renal morphogenesis. To date, the effects of SHH on fibroblast growth factors (Fgfs) in embryonic kidney development remain unclear. In the present study, explants of BALB/c mouse embryonic kidney tissues were used to investigate the role of exogenous SHH on Fgf8 and Fgf10 expression levels ex vivo. Ureteric bud branches and epithelial metanephric derivatives were used to determine the renal morphogenesis with Dolichos biflorus agglutinin or hematoxylin­eosin staining. mRNA expression levels were determined using reverse transcription­quantitative polymerase chain reaction, while the protein expression levels were examined using immunohistochemistry and western blot analysis. During the initial stages of metanephric development, low levels of SHH, Fgf8, and Fgf10 expression were observed, which were found to increase significantly during more advanced stages of metanephric development. In addition, exogenous SHH protein treatment increased the number of ureteric bud branches and enhanced the formation of nephrons. Exogenous SHH reduced the Fgf8 mRNA and protein expression levels, whereas cyclopamine (an SHH­smoothened receptor inhibitor) interfered with SHH­mediated downregulation of Fgf8 expression. By contrast, exogenous SHH protein was not found to modulate Fgf10 mRNA and protein expression levels. In conclusion, these results indicate that the modulatory effects of SHH on BALB/c mouse metanephric explant cultures may involve the regulation of Fgf8 expression but not Fgf10 expression, which provides evidence for the functional role of Fgf proteins in renal morphogenesis.

Plasma 15-F2t-isoprostane in idiopathic pulmonary arterial hypertension.

BACKGROUND: 15-F2t-isoprostane (15-F2t-IsoP), a prostaglandin F2-like compound, is widely recognized as a biomarker of chronic heart failure. This study investigated the potential role and prognostic significance of plasma 15-F2t-IsoP in patients with idiopathic pulmonary arterial hypertension (IPAH). METHODS: Plasma 15-F2t-IsoP concentrations were determined in 80 consecutive IPAH patients at the time of their first right heart catheterization, and monitored for 30±12 months. The expression of 15-F2t-IsoP protein in autopsy lung samples was determined by immunohistochemical staining. RESULTS: Plasma 15-F2t-IsoP concentrations were significantly increased in patients with IPAH compared with healthy controls (91 pg/ml vs. 30 pg/ml, respectively; P<0.001). Patients with baseline 15-F2t-IsoP concentrations≥97 pg/ml had a significantly lower survival rate than those with lower baseline concentrations (P<0.001). During follow-up, 15-F2t-IsoP concentrations in survivors decreased, whereas concentrations in non-surviving patients increased further (P<0.05). Elevated concentrations of 15-F2t-IsoP were correlated with a severity of WHO functional class, lower 6-minute walking distance and mixed venous oxygen saturation, higher mean right atrial pressure and brain natriuretic peptide. Multivariate analysis revealed that the plasma 15-F2t-IsoP concentration was an independent factor associated with mortality. Histological studies showed that the expression of 15-F2t-IsoP was up-regulated in remodeled pulmonary vessels. CONCLUSIONS: An elevated plasma 15-F2t-IsoP concentration and a further increase during follow-up may be a risk factor for higher mortality in patients with IPAH.

The phosphodiesterase-5 inhibitor vardenafil reduces oxidative stress while reversing pulmonary arterial hypertension.

AIMS: Oxidative stress is implicated in the pathology of pulmonary arterial hypertension (PAH). Previously, we demonstrated that vardenafil, a phosphodiesterase-5 inhibitor, has potential as therapy for PAH, although the mechanism remained uncharacterized. Here, we aimed to determine baseline levels of oxidative stress in PAH and investigate whether vardenafil affects oxidative stress levels while improving PAH. METHODS AND RESULTS: Sprague-Dawley rats with monocrotaline-induced PAH were administered oral vardenafil 1 mg kg⁻¹ day⁻¹ for 21 days (n = 12). Treatment-naive patients (n = 15) with PAH were treated with vardenafil 5 mg twice daily for 3 months. Haemodynamic data and plasma levels of nitrate/nitrite and products of oxidative damage were determined in rats and patients. Histopathology, immunohistochemistry, and assessments of oxidative/anti-oxidative enzyme expression were performed in rat lung tissue. Compared with baseline (patients) or untreated controls (rats), vardenafil significantly reduced pulmonary vascular resistance and increased cardiac output (CO). In rats, vardenafil suppressed proliferation and enhanced apoptosis of pulmonary artery smooth muscle cells, attenuating small pulmonary artery remodelling, and right ventricular hypertrophy. Vardenafil significantly reduced levels of oxidative stress biomarkers, such as 8-iso-prostaglandin-F2α and 3-nitrotyrosine, and significantly increased nitric oxide (NO) levels in rats and patients. Furthermore, vardenafil significantly increased endothelial NO synthase expression and superoxide dismutase activity, and down-regulated nicotinamide adenine dinucleotide phosphate oxidase expression in rat lung tissue. CONCLUSION: Vardenafil reduces oxidative stress in rats and humans while improving PAH, warranting investigation of oxidative stress pathways as targets for PAH therapy.