Evaluation of combined photodynamic inactivation and no releasing chitosan nanoparticles on cutaneous leishmaniasis

Abstract

Cutaneous leishmaniasis (CL) is a chronic disease developed by parasites of the genus Leishmania that promotes destructive lesions. The available treatments are limited because of side effects, resistance and toxicity. New strategies against CL have been studied such as Photodynamic inactivation (PDI) and exogenous NO donors. The aim of this work was to explore the effects of methylene blue (MB)-mediated PDI in association with encapsulated NO donors in chitosan nanoparticles (NPNO) on Leishmania amazonensis. NPNOs were tested in vitro with L. amazonensis transgenic line expressing luciferase at increasing concentrations (25-200μM) and inhibitory concentrations (IC50 and IC90 ) were calculated. Based on inhibitory concentrations results, twelve BALB/c mice were infected in the left footpad and randomly assigned to experimental groups (n=4): Control (non-treated), G1 (two PDI sessions), G2 (two PDI sessions and 80 μM of NPNO, immediately after PDI) and G3 (only 80 μM NPNO). PDI was performed using a red LED (λ= 660±22 nm) at 150 J/cm² fluence and MB at 100 μM. Parasite burden was obtained by bioluminescence every day, in the first 96 h and for the next 4 weeks, once a week. Test groups presented significant reduction in parasite load compared to control during all experimental period. In the first 24 h after treatments, parasite burden was significant lower for G2. After 96 h, all test groups were similar. Following 4 weeks, statistically significant differences were noticed when test groups were compared to control but parasite burden was similar among all treated groups. Under conditions used in this study, our results show that NPNOs were not able to sustain the parasite killing promoted by MB-mediated PDI on CL induced in mice 24 h after treatments.