Ação imunomodulatória da cloroquina em células mononucleares do sangue periférico de pacientes com diabetes mellitus tipo 1

Abstract

Type 1 Diabetes mellitus (T1DM) is an autoimmune disease, characterized by chronic hyperglycemia, which generates an inflammatory process that results in micro and macrovascular complications. The treatment of T1DM is directed towards glycemic control through the use of insulin, however, it has not been sufficient to control or reduce the inflammatory process. In this sense, adjuvant therapies combining anti-inflammatory drugs and nanoparticles (NPs) have emerged as a strategy to prevent and / or delay the complications associated with this disease. Chloroquine (CQ), because it has anti-inflammatory properties, becomes one of the alternatives of adjuvant treatment for diabetes. Thus, the objective of this study was to evaluate the use of chloroquine, incorporated or not incorporated in nanoparticles of polylactic acid (PLA), in peripheral blood mononuclear cells (PBMC) of patients with T1DM, in order to observe its immunomodulatory effect on IL-1β cytokines , IL12, IFN-γ, TNF-α and IL-10. Twenty five normoglycemic individuals and Twenty five patients with T1DM were selected, both aged 10 to 16 years. Glycemic control was assessed using glycated hemoglobin (HbA1c) and serum glucose concentration. The viability of the T1DM group PBMCs was assessed by the MTT test by exposure to CQ and CQ incorporated into nanoparticles (CQ NPs) in concentrations of 200, 100, 50, 25, 10 and 5 µM. The CQ concentration of 10µM was chosen and the PBMCs of T1DM patients were submitted to three treatment conditions: untreated (NT), treated with chloroquine (CQ) and treated with chloroquine incorporated in nanoparticles (CQ NPs). The cells were incubated for a period of 48 hours and at 24 hour intervals the mRNA expressions of the cytokines IL1B, IFNG, TNFA, IL12 and IL10 were determined by relative quantification in real time PCR. IL1B expression was reduced in cells treated with CQ within 48h (p <0.001) and in CQ NPs after 24h of treatment (p <0.05); the expression of the IFNG and IL12 genes was significantly reduced (p <0.001) in cells treated with CQ and CQ NPs at 24 and 48h compared to NT; TNFA showed a significant reduction only after 48 hours of treatment with CQ and CQ NPs (p <0.0001) and IL10 expression showed a significant reduction only after 24 h of treatment with CQ and CQ NPs (p <0.002). Thus, it was possible to demonstrate that CQ has anti-inflammatory activity in the primary cells of patients with T1DM, which may represent an alternative and adjuvant anti-inflammatory therapy to prevent diabetes complications. In addition, CQ nanoparticles represent a promising innovation, offering a safe vehicle for their intracellular action and better control in prolonged treatments.