Preparation, characterization and potential therapeutic effect in Arterial Hypertension treatment of Anandamide/Polycaprolactone nanoparticles

CARLOS GAMARRA-LUQUES; DIEGO KASSUHA; VIRNA MARTÍN GIMENÉZ; LUCIA FUENTES; WALTER MANUCHA; LUCIANA MAZZEI; JOSEP ESTEVE ROMERO

Kyoto

Simposio; 49th International Symposium on High Performance Liquid Phase Separations and Related Techniques; 2019

Kyoto University at Katsura

Introduction: Anandamide (N-arachidonoil ethanolamine, AEA) was the first endogenous ligand of central (CB1) cannabinoid receptors. It has demonstrated several effects on mammalian cells, including nitric oxide-mediated hypotension. However, it has several disadvantages: as it is a fatty acid derivative, it shows poor aqueous solubility and stability, and also has undesirable psychoactive effects. In this sense, the use of nanotechnology to achieve a prolonged and localized release of AEA as well as to improve its aqueous solubility and stability by reducing its adverse effects would offer an interesting approach and a great challenge.Objective: To Prepare and characterize AEA loaded polycaprolactone (PCL) nanoparticles (Nps), and to evaluate its biocaptation and activity in renal cells and its stability.Materials and Methods: PCL nanoparticles loaded with AEA were obtained by electrospraying technique. The morphology and particle size distribution were determined by means of SEM. In addition, thermal properties (DSC, TGA), interactions between drug and polymer and physical and chemical stability (FTIR) at 4, 25 and 40 ˚C were studied. Additionally, Western blot measurements of inducible isoform of Nitric Oxide synthase (iNOS) expression, as well as Nitric Oxide (NO) detection and Na+/K+ ATPase activity were made in duplicate with six cultures of human proximal tubule cells (HK2): control; PCL Nps (without AEA); PCL/AEA Nps (1 μM and 10 μM); free AEA (1 μM and 10 μM).. Results: SEM images evidenced ovoid particles distributed in a range of 100-900 nm with a predominance of a population of sizes between 200 and 400 nm. DSC curves showed endothermic fusion peaks of AEA (40 ˚C) and PCL (60 ˚C). The NPs exhibited a fusion endotherm of 55.85 ˚C, which means that thermal stability was increased with respect to the pure drug. FTIR demonstrated the existence of intermolecular interactions due to the displacements of the C=O, OH and NH bands. The obtained NPs remained stable under all the conditions studied at least for 120 days. iNOS expression and NO concentration were increased with the AEA treatment with respect to the controls. Instead, Na+/K+ ATPase activity was declined. In all cases, differences were observed between encapsulated and free AEA.Conclusions: The electrospraying technique allowed the obtaining of NPs with attractive physicochemical properties and sufficient stability. Those NPs were able to supply AEA in a controlled manner and decrease Na+/K+ ATPase activity at the renal level through a mechanism mediated by iNOs and NO. This finding would allow us to suggest a potential natriuretic/diuretic effect of PCL/AEA NPs which would be promising for the treatment of arterial hypertension.