Figure 5. The activation of NADPH oxidase and the impaired relaxation to ACh induced by visfatin in rat microvessels relies on Nampt activity.
(A) NADPH oxidase activity was measured in cultured human umbilical vein endothelial cells (HUVEC) stimulated for 20–30 min with visfatin (50 ng/mL), the Nampt inhibitor APO866 (10 µmol/L) or both compounds together, as well as with the product of the Nampt reaction NMN (10 and 100 µmol/L). Results are expressed as mean ± SEM of 5 experiments performed in triplicate. *P<0.05 vs control cultures; †P<0.05 vs visfatin (B) Effect of visfatin (50 ng/mL) and APO866 (10 µmol/L), alone or in combination, on NADPH oxidase activity in rat microvascular preparations. Results are expressed as mean ± SEM of 8 segments obtained from 4 animals. *P<0.05 vs control cultures; †P<0.05 vs visfatin. (C) Rat microvessels were pre-incubated with visfatin (50 ng/mL), alone or in the presence of APO866 (10 nmol/L to 10 µmol/L), subsequently contracted with 1 µmol/L NA and then exposed to increasing concentrations of ACh (100 pmol/L to 3 µmol/L). Results are expressed as mean ± SEM of 58 segments obtained from 11 animals. *P<0.05 vs control curve; †P<0.05 vs visfatin alone. (D) Correlation between the APO866 concentrations used for pre-incubation and the pD2 values for ACh. (E) The vessels were pre-incubated with NMN (100 nmol/L to 1 mmol/L), contracted with 1 µmol/L NA and then relaxed with cumulative concentrations of ACh (100 pmol/L to 3 µmol/L). Results represent the mean ± SEM of 32 segments obtained from 5 animals. *P<0.05 vs control curve. (F) Correlation between the NMN concentrations used for pre-incubation and the pD2 values for ACh.