Medida de óxido nítrico por amperometria induzida pela N-saliciloiltriptamina em células endoteliais recém dispersas

Abstract

The vascular endothelium is responsible for maintaining homeostasis in the cardiovascular system by controlling blood pressure by releasing relaxing and constrictors endothelium - derived factors. Among the relaxing factors, NO (nitric oxide) presents a majo r role in the control of vascular tone through activation of the sCG/cGMP leading to the relaxation of VSMC (vascular smooth muscle cell). However, a simple hemodynamic phenomenon may be the result of summation other endothelium - derived relaxing mechanisms acting in concert, it is so great importance to measure NO to understand the role of this molecule in various biological processes. Therefore, the objective of this work was to implement the method of measurement of NO by amperometry at newly scattered en dothelial cells of aorta and evaluate the amount of NO release d by STP, a derivative of benzoyltryptamine s where previous studies have shown tha t it was capable of causing relaxation e ndothelium - dependent in mesenteric artery rings and this effect was rela ted to the participation of the NO/sCG/ cGMP. The aorta of Wistar rats (250 - 300 kg) was removed and immersed in Hank's solution whose endothelial cells (ECs) were isolated by means of a mechanical scraping. An aliquot of the ECs of suspension was sent to fl ow cytometry for the study of identification and cell viability (over 75%). Then, calibration was performed using a microsensors of CuCl 2 solution and a spontaneous NO donor, SNAP. Finally, increasing concentrations were added to STP (1; 10 and 100 μ M) for performing the measurement of NO by amperometry. The results of the flow cytometry was 90% of the cells present in the suspension were viable and 79% were ECs. The calibration of microsensors generated a linear regression graph (R 2 = 0.998756), indicating that this procedure produced an excellent linear calib ration probe NO. Finally the STP was able to significantly increase (P <0.05) the concentration of NO from endothelial c ells isolated suspension of aorta. Therefore, we conclude that this measurement o f NO by amperometry technique allows to detect small quantities of this molecule in solution is useful in single cell preparations, and the ability to assess the local production of nitric oxide, facilitating the operation of this mediator role in physiolo gical processes or pathophysiological