IN VITRO TOLERANCE AFTER INCUBATION WITH NITROGLYCERIN (NTG) IS DUE TO INCREASED REACTIVE OXIGEN SPECIES IN THE ENDOTHELIAL CELLS

Abstract

NTG may cause tolerance during long‐term administration, which is its major therapeutic limitation. However, the mechanisms underlying NTG‐induced tolerance and the contribution of endothelial cells (EC) remain unclear. We have verified that the endothelium modulates the vasodilatation induced by NTG in rat aorta. This work aimed to study the effects of nitric oxide (NO) and reactive oxygen species (ROS) on NTG‐induced relaxation in rat aorta and in isolated EC in order to investigate the cause of the in vitro tolerance. Relaxation curves for NTG were constructed in intact endothelium (E+) aortic rings. It was determined the efficacy (ME) and potency (pD2) of NTG. L‐NAME reduced the pD2 (from 8.70±0.13 to pD2 7.83±0.15) and ME from 102.7±2.7%, n=10 to 69.8± 3.2%, n=5 (p<0.001). To study the tolerance in vitro, the E+ arteries were incubated for 5 min with NTG in the concentration that produced the ME: 100 μM. The ME and pD2 were reduced (from 101.9±1.6%; n=5 to 54.7±3.6%, n=5 p<0.001; and from 8.88±0.20 to 7.63±0.04; n=5, p<0.05), respectively. Following 5 min incubation, NTG enhanced the fluorescence intensity units (U) of superoxide (O2−) concentration by flow citometry (from basal 2.19±0.58U to 54.15±1.49U, p<0.001). NO concentration was not changed in the EC. In conclusion, the in vitro tolerance is induced by 5 min incubation with NTG with increased production of superoxide in aortic EC. Supported by FAPESP and CNPq.