Influence of Extracellular Traps (ETs) on the Differentiation of TCD4 Cell Profiles and Macrophages in Human Autologous Culture

Abstract

Background: The formation of extracellular traps (ETs) as a microbicidal functional mechanism of various leukocytes has taken on importance today and they are implicated in the pathogenesis of various diseases and have even been involved in the current pandemic. Objectives: Study ETs generated in vitro from healthy human blood leukocytes against different stimuli (LPS, fMLP) and their influence on different T cells profiles and monocyte-macrophages, in autologous cell cultures. Methods: Heparinized human blood samples were collected with ethical consent. ETs generation was performed by stimulation with LPS and fMLP. Subsequently they were isolated.  ETs influence on cell profile differentiation was performed in samples without stimulation, with OVA addition samples, and OVA-ETs addition samples. This assay was observed through immunofluorescence (IF) labeling of molecules of T CD4 profile; Th17 and innate lymphoid cells 3 ILC3 by RORɣ; activation status of T cells by CD45RO; and M1 macrophage profile by iNOS. Results: Significant CD4 and CD45RO positive cells percentage were observed between paired control samples and OVA addition samples (p <0.05), between paired control samples and OVA-ETs addition samples (p <0.05). In an independent experiment, significant differences were observed between OVA addition samples vs. OVA-ETs addition samples (p <0.05). At 72 h of culture, no significant differences were found between the paired samples in any case. There were no significant differences between paired control samples and OVA addition samples or OVA-ETs samples, neither 24 and 72 h of culture in RORɣ positive cells percentage or iNOS positive cells percentage. Conclusions: Influence of ETs on T cell activation was observed and components of autologous ETs did not elicit classical activation of M1 macrophages.