Avaliação In Vitro Da Atividade Antileishmania E Imunomoduladora Do Galato De Octila Sobre Formas Amastigotas De Leishmania (Leishmania) Infantum

Abstract

Neglected tropical diseases (NTDs) affect more than one billion people, with a significant impact in impoverished regions such as Northeastern Brazil, especially the state of Piauí. Among these diseases, visceral leishmaniasis (VL) stands out due to its severity, lethal effects, and socioeconomic burden. Current treatment, based on first- and second-line drugs, is limited by high toxicity, adverse effects, and growing parasite resistance. In this context, octyl gallate (OG), a derivative of gallic acid with antioxidant and antiparasitic properties, emerges as a promising alternative against Leishmania spp. Therefore, the aim of this study was to evaluate the activity of OG on amastigote forms of Leishmania (Leishmania) infantum internalized in macrophages, as well as to investigate its effects on immune mediators. RAW 267.4 cells were infected with L. infantum promastigotes and treated with different concentrations of OG for periods of 24 and 72 hours. Analysis was performed by optical microscopy, evaluating the percentage of infected cells and the number of amastigotes per macrophage. OG demonstrated a significant anti-leishmanial effect, with progressive inhibition of infection at all tested concentrations. The EC50 was 3.87 μM (24 h) and 3.04 μM (72 h), indicating greater efficacy of OG at 72 hours. To assess the immunomodulatory effects induced by OG, the supernatants

from the infection assays were analyzed by sandwich ELISA to quantify the production of pro- inflammatory cytokines (TNF-α and IL-12) and anti-inflammatory cytokines (IL-10 and IL-6)

produced by macrophages infected with L. infantum. The results showed that OG induced a Th1 immune response, with a significant increase in TNF-α and IL-12 production, while IL-10 levels were significantly reduced and IL-6 levels remained unchanged under the evaluated conditions. These data indicate the immunomodulatory potential of OG, promoting an effective pro-inflammatory environment against the parasite. Nitric oxide (NO) production by macrophages infected with L. infantum and treated with OG was indirectly evaluated by measuring nitrite (NO2−) levels. After 24 and 72 hours of incubation, the culture supernatants were analyzed. After 72 hours, the 25 μM concentration was the only one to show a significant effect compared to the control. At 24 hours, an increase in nitrite was also observed at the 25 μM concentration. It can thus be concluded that OG is effective against L. infantum in macrophages, efficiently inhibiting infection with high potency even at low concentrations. In addition to its antiparasitic action, OG stimulates a Th1 immune response, increasing TNF-α and IL-12, reducing IL-10, and activating macrophage defense mechanisms by promoting NO production. Therefore, OG is a promising candidate for treatments against VL.