MICROBIOTA, IGG AND CD64+ MACROPHAGES:THEIR INTERACTIONS IN THE COLON LAMINA PROPRIA

Congreso; REUNIÓN CONJUNTA DE SOCIEDADES DE BIOCIENCIAS; 2017

Intestinal macrophages (MΦs) identified by CD11b, F4/80 andCD64 markers play homeostatic roles in the colonic environment. Toevaluate frequency and localization of these cells, whole mount colonsections were stained with fluorescent anti-CD64 and anti-CD11bantibodies. As these MΦs can uptake immune complexes (ICs), weused ex vivo and in vitro strategies to evaluate the uptake of IgG-ICsand stimulatory activity during inflammatory conditions. Ex vivo, weobtained colonic lamina propria MΦs from control and colitis mice byenzymatic digestion and cell sorting using the F4/80 marker. MΦswere stimulated 1 h at 37°C with ICs and the internalized IgG wasevaluated by Western blot. In vitro, bone marrow-derived MΦs werecultured with conditioned medium obtained out of colonic explantcultures from control or colitis mice; in the later condition a significantincrement in Ly6c and F4/80 expression was found (p<0.05),possibly due to the inflammatory mediators released by colitis tissue.When these MΦs were stimulated with IgG-ICs, we found asignificant decrease in geometric mean fluorescence intensity forCD64 (p<0.05), Ly6c (p<0.05) and F4/80 (p=0.008) either with conditionedmedium from control or colitis mice. As we previously foundthat the opsonization of fecal bacteria increases significantly duringcolitis development, bone marrow-derived MΦs were stimulated for48 h with fecal bacteria opsonized with IgG and IgA and sorted fromcontrol or colitis mice to evaluate the proinflammatory potential ofluminal ICs; NO, IL-6 and IL-10 were evaluated in supernatants bycolorimetric assays. Our results show that the CD64+ CD11b+ MΦsrepresent an abundant population in the colonic mucosa, localizedaround the cripts in narrow contact with the epithelial layer. Inflammatoryenvironment modulated the phenotype of MΦs meanwhilethe ICs down-modulated mainly CD64 and F4/80, suggesting acrosstalk between these molecules.Keywords: MΦs, colonic mucosa, IgG-ICs, microbiota