Role of Cox Pathway in Human Monocytes and Monocyte‐Derived Dendritic Cells infected by Trypanosoma cruzi
Authors: Lonien Sandra Cristina Heim , de Freitas Rafael Carvalho , Suzukawa Helena Tiemi , dos Santos Fernando Fernandes , Lovo‐Martins Maria Isabel , Silveira Guilherme Ferreira , Wowk Pryscilla Fanini , Bordignon Juliano , Pinge‐Filho Phileno 
Published: November 2023

Studies have shown that nonsteroidal anti‐inflammatory drugs (NSAIDs) modulate innate inflammatory response and inhibits the invasion of T. cruzi into macrophages controlling Chagas disease progression. PGE2, together with nitric oxide (NO) and TNF‐α, participate in a complex circuit that controls immune responses in T. cruzi infection. However, the involvement of cyclooxygenase‐mediated PGE2 production in the entry of T. cruzi into human monocytes and dendritic cells has not been elucidated. The aim of this study is to explore the effect of inhibition of cyclooxygenase 1 and 2 on the internalization of T. cruzi (Y strain) and regulation of immune response in human monocytes and monocyte‐derived dendritic cells (moDCs). Blood samples were obtained from healthy volunteers. (Approved by the Ethical Committee in Research Involving Human Subjects of State University of Londrina: 5491/2012 and CONEP: 5231. Mononuclear cells were separated by density gradient centrifugation using lymphocyte separation medium (Lonza, Walkersville, MD). Adherent purified CD14+ (monocytes), used to derived dendritic cells (moDCs) after cultured for 6 to 7 days in RPMI medium containing interleukin‐4 (100 ng/mL) and granulocyte‐monocyte colony‐stimulating factor (50 ng/mL), 10% fetal bovine serum, 100 IU/μg/ml penicillin/streptomycin, and 2 mM L‐glutamine (Gibco‐BRL, Grand Island, NY). Both cell types were pretreated with aspirin (ASA) or celecoxib (CEL) (0.312 – 1.25 mM) one hour before T. cruzi infection (5:1). The cells were washed, fixed with methanol fixative, stained with Giemsa (Merck) stain, and observed with a light microscope for determination of internalization index. Production of NO was determined by Griess reaction. Levels of IL‐12p70 and IL‐10 in culture supernatants were measured by commercial ELISA kits (eBioscience, SanDiego, CA) or cytometric bead assay human inflammatory kit (Becton Dickinson, CA, USA), according to the manufacturer's instructions. We showed that NO, PGE2, IL‐10 and IL‐12 production increased significantly in the monocytes cultures following exposure to viable trypomastigotes forms derived from the supernatant of T. cruzi‐infected LLC‐Mk2 culture cells. ASA and CEL increased synthesis of NO and IL‐12 in a dose‐dependent manner, whereas celecoxib increased both IL‐10 and IL‐12. Pharmacological blockade of COX‐1 (ASA) and COX‐2 (CEL) caused inhibition of T. cruzi infection in both monocytes and moDCs. In contrast to monocytes, this inhibition was independent of NO production in moDCs cultures. Unlike what happened with monocytes in all tested concentrations of the drugs, there was no significant effect of ASA on internalization of T. cruzi by dendritic cells; however, the effect of C was dose‐dependent. Altogether, these results support the idea that the COX pathway plays a role in the process of parasite invasion in human monocytes and moDCs, and an effect for NSAIDS therapies on the innate inflammatory response to T. cruzi infection.Support or Funding InformationCAPES, CNPQ, Fundação Araucária