CLOSTRIDIOIDES DIFFICILE UPTAKE BY HUMAN MACROPHAGES OCCURS IN A SLAMF1-INDEPENDENT MANNER

BARBERO AM; HERNÁNDEZ DEL PINO RE; ESPAÑOL LA; MORRO LS; MACHAIN M; FUENTES F; BARRIONUEVO P; PASQUINELLI V

VIRTUAL

Congreso; Reunión Conjunta SAIC. SAI. AAFE. NANOMED-AR; 2021

SAIC, SAI, AAFE Y NANOMED-AR

C. difficile (CD), a Gram+ spore-forming anaerobic bacteria, is the major cause of nosocomial infectious diarrhea that generally develops after antibiotic treatment. SLAMF1 (Signaling Lymphocytic Activation Molecule) is expressed on macrophages and triggers not only phagosome-related functions, but can also recognize and internalize different pathogens. CD components regulate macrophages functions and both toxins and spores invade host cells. However, the endocytosis of the bacteria has not been explored. Here, we address the internalization of CD and the role of SLAMF1 during this process.Monocyte-derived macrophages from healthy donors were cultured in the presence or absence of CD (NAP1/BI/027 strain) inactivated by heat or formalin treatment (CDH or CDF). For some experiments CD was coupled with FITC and an agonistic antibody for SLAMF1 was added to the cell culture.Our results show that SLAMF1 expression is not modulated on human macrophages surface by either CDH or CDF, even in the presence of increasing amounts of bacteria as shown by flow cytometry. Biochemical assays were conducted and SLAMF1 detection with a specific antibody indicated that there is no interaction between SLAMF1 and CDH or CDF. Nevertheless, we did detect interaction of macrophages with CD. SLAMF1 is also a costimulatory molecule, but SLAMF1 costimulation through an agonistic antibody had no effect on the CD-macrophage interaction as measured by flow cytometry and fluorescence microscopy. Moreover, most of the inter- acting macrophages were SLAMF1 negative. Finally, we confirmed intracellular localization of C. difficile in macrophages and a partial colocalization with LAMP2 by confocal microscopy.In conclusion, SLAMF1 does not promote the interaction between human macrophages and C. difficile and does not participate in the internalization process of the bacterium. Further studies are needed to elucidate the molecules involved in macrophage entry mechanisms of C. difficile.