Efeito modulatório in vitro da quetiapina não-metabolizada na citotoxicidade e ativação inflamatória de células brancas e na formação de armadilhas extracelulares de neutrófilos

Abstract

Quetiapine (QUE) is a highly complex drug that, due to its different forms of interaction with receptors, has great clinical applicability. Although widely used, QUE brings with it a range of side effects that include, in addition to disorders of the metabolic pathway, changes in levels of the immune system. Quickly metabolised by the liver, it is transformed into its main active metabolite, norquetiapine, being then the clearance of QUE only 5%. This percentage may increase in elderly people, with liver failure or through interaction with other drugs. Thus, non-metabolized quetiapine (nmQUE) may be responsible for inducing peripheral immune changes, especially low-grade chronic inflammation. Therefor, our objective was to evaluate in vitro the potential cytotoxicity and modulatory effect of nmQUE on inflammatory activation and in the formation of extracellular neutrophil traps (NETs). For this, cellular experiments were carried out, using a commercial macrophage cell line (RAW 264.7), in addition to leukocytes and neutrophils, obtained through blood collection from volunteers. The cells were cultured under ideal conditions according to standardisation until the concentration of 1x105 cells/mL was obtained. Then, macrophages, leukocytes and neutrophils were treated and activated, or not, with a mitogenic agent, phytohemagglutinin (PHA). After 24 hours, they were treated with different concentrations of QUE (25, 50, 100, 200 and 400 μg/L). Given at 24 hours of treatment, the cell viability and cytotoxicity of all cells were evaluated using the 3- [4,5-dimethylthiazol-2-yl] -2,5-diphenyltetrazolium assay (MTT). With the result of these tests, the concentration of choice for QUE to be used in the following analyses was defined: 100 μg/L. Thus, after 72 hours of treatment, cell proliferation, oxidative molecules and gene and protein expression (via Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) of pro-inflammatory and anti-inflammatory cytokines in RAW 264.7 were evaluated. In addition, to assess the effect of nmQUE on the formation of NETs, neutrophils were previously activated, or not, by exposure to yeast cells for 2 hours, followed by exposure to QUE for an additional 2 hours, then fixed and stained, followed by images capture and evaluations consistent with the experiment.The results show the high capacity of nmQUE to interact with immune cells, not showing cytotoxicity, but modifying its inflammatory profile according to its initial state, thus corroborating it with a pro and anti-inflammatory pharmacological effect. In relation to the formation of NETs, neutrophils were surprisingly induced, with nmQUE being responsible for hyper-formation of NETs. With these results, it is concluded that nmQUE exerts an influence on the pathways of the immune system, being possibly responsible for the existence of morbidities and side effects, which represents the major problem of antipsychotic therapy.