Ex Vivo and In Silico Approaches of Tracheal Relaxation through Calcium Channel Blockade of 6-Aminoflavone and Its Toxicological Studies in Murine Models.

Asthma is a condition in which a person's airways become inflamed, narrowed, and produce greater amounts of mucus than normal. It can cause shortness of breath, chest pain, coughing, or wheezing. In some cases, symptoms may be exacerbated. Thus, the current study was designed to determine the mechanism of action of 6-aminoflavone (6-NHF) in ex vivo experiments, as well as to determine its toxicity in acute and sub-chronic murine models. Tissues were pre-incubated with 6-NHF, and concentration-response curves to carbachol-induced contraction were constructed. Therefore, tracheal rings pre-treated with glibenclamide, 2-aminopyridine, or isoproterenol were contracted with carbachol (1 µM), then 6-NHF relaxation curves were obtained. In other sets of experiments, to explore the calcium channel role in the 6-NHF relaxant action, tissues were contracted with KCl (80 mM), and 6-NHF was cumulatively added to induce relaxation. On the other hand, tissues were pre-incubated with the test sample, and after that, CaCl concentration-response curves were developed. In this context, 6-NHF induced significant relaxation in ex vivo assays, and the effect showed a non-competitive antagonism pattern. In addition, 6-NHF significantly relaxed the contraction induced by KCl and CaCl, suggesting a potential calcium channel blockade, which was corroborated by in silico molecular docking that was used to approximate the mode of interaction with the L-type Ca channel, where 6-NHF showed lower affinity energy when compared with nifedipine. Finally, toxicological studies revealed that 6-NHF possesses pharmacological safety, since it did not produce any toxic effect in both acute and sub-acute murine models. In conclusion, 6-aminoflavone exerted significant relaxation through calcium channel blockade, and the compound seems to be safe.