MURINE MACROPHAGES SURFACE N-GLYCANS PARTICIPATE IN YopP-MEDIATED IMMUNOPATHOGENESIS OF Yersinia enterocolitica INFECTIONS

GARAY, JUAN A.; SILVA, JUAN E.; VARAS J.; FUNES, SAMANTA C.; JOFRÉ, BRENDA L.; DI GENARO, MARÍA S.; DAVICINO, ROBERTO C.

Mar del Plata

Congreso; LXX REUNIÓN ANUAL DE LA SOCIEDAD ARGENTINA DE INMUNOLOGÍA (SAI) & 3ER CONGRESO FRANCO-ARGENTINO DE INMUNOLOGÍA (FAIC); 2022

ASOCHIN

Yersinia enterocolitica (Ye) outer membrane protein P (YopP) is able to manipulate macrophages (MΦ) by inducing apoptosis and suppressing signaling molecules production. Although YopP has been shown to interact in a carbohydrate-dependent manner with host proteins, its interaction with glycans remains unclear. The aim of our work is to explore the influence of MΦ surface N-glycans on the immunomodulatory effects triggered by YopP. Murine peritoneal MΦ were obtained by intraperitoneal lavage, plated at a 1x106 cells/mL and purified by incubation for 2 h at 37°C in DMEM medium in an atmosphere with 5% CO2. Then, MΦ were incubated with 5 IU/mL of PNGase F in DMEM medium without Fetal Bovine Serum for 4 h (MΦd) and cultured overnight in supplemented DMEM medium. Subsequently, MΦ were infected with Ye serotype 0:8 (pYV+, WA 314) (Ye wt) or with Ye WA-314 deficient in YopP (pYV+, WA-CpYVNalrKanr) (Ye ∆yopP) at MOI 10:1. Then, MTT, lactate dehydrogenase (LDH), Griess and Urease assays were performed. Apoptosis was evaluated by flow cytometry using Annexin V-propidium iodide, and TNF-α was measured by ELISA. Our results shows that N-deglycosylation does not affect viability. Besides, we observed no significant apoptosis in Ye wt-infected MΦd compared to uninfected counterpart, suggesting a critical impact of N-glycans in the wellknown triggering role of Ye on apoptosis. On the other hand, we showed that YopP has a dual effect: in MΦ inhibits NO production (p