Human Polymorphonuclear Neutrophil Phenotypes Generated in vitro

Abstract

Background: There are a variety of polymorphonuclear neutrophil phenotypes described in different species and health or disease situations.  Objective: Study human neutrophil phenotypes generated in vitro. Methods: Heparinized human blood samples were collected with ethical consent. Polymorphonuclear neutrophils purification and autologous cultures was performed. Neutrophil stimulation was performed with LPS, fMLP or OVA. Immunofluorescence was applied.  Results: “Polymorphonuclear neutrophil-antigen presenting cell” profile was generated in vitro, expressing CD80, CD86 and HLA-DR molecules. Immunofluorescence analysis show: CD80 expression, significant differences between CTFT control and CTFT fMLP (p<0.05), CTFT control and CTFT OVA (p<0.0001). CD86 expression, significant differences between CTFT control and CTFT fMLP (p<0.05), CTFT control and CTFT LPS (p<0.05), CTFT control and CTFT OVA (p<0.0001). HLA-DR expression, significant differences between CTFT control and CTFT LPS (p<0.05). About “Polymorphonuclear neutrophil-CD4-CD45RO” profile, analysis show: CD4 expression, significant differences between CTFT control and CTFT fMLP (p<0.05). CD45RO expression, no significant differences. “Polymorphonuclear neutrophil-antigen presenting cell” phenotype, released NETs with CD80, CD86 at 30 minutes: paired control samples (7.4%), stimulated with LPS (12.69%), fMLP (16.67%) and OVA (18.47%). HLA-DR expression in NETs, at 30 minutes, in paired control samples (0%), stimulated with LPS (16.17%). At 17 hs, in paired control samples (0%), with OVA stimulation (4.54%). “Polymorphonuclear neutrophil-CD4-CD45RO” phenotype, released NETs expressing CD4 and C45RO molecules. At 30 minutes, in paired control samples (0%), stimulated with LPS (7.67%), fMLP (6.38%) and OVA (0%). Conclusions: Molecules expressed by phenotypes can play a relevant role by influencing cellular microenvironment and can be taken into account as possible therapeutic targets.