Extracellular vesicles of porphyromonas gingivalis impair trophoblast cell interaction with immune Cells.

B LARA, G CALO, M SASSOT, L GLIOSCA, I LOUREIRO, S GORI, L CASTAGNOLA, D PAPARINI, R RAMHORST, D VOTA, C PÉREZ LEIRÓS, V HAUK

San LUis

Congreso; Reunión Anual de la Sociedad Argentina de inmunología (SAI); 2023

Sociedad Argentina de inmunología (SAI)

124 (171) EXTRACELLULAR VESICLES OF PORPHYROMONASGINGIVALIS IMPAIR TROPHOBLAST CELL INTERACTION WITH IMMUNECELLSBrenda Lara1, Guillermina Calo1, Matías Sassot1, Laura Gliosca2, Iñaki Loureiro1, SoledadGori1, Lara Castagnola1, Daniel Paparini1, Rosanna Ramhorst1, Daiana Vota1, Claudia PérezLeirós1, Vanesa Hauk11 Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Departamento deQuímica Biológica, Laboratorio de Inmunofarmacología. Buenos Aires, Argentina. CONICET,Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN),Buenos Aires, Argentina. 2 Universidad de Buenos Aires, Facultad de Odontología. Laboratoriode Diagnóstico Microbiológico, Cátedra de Microbiología. Buenos Aires, Argentina.Periodontitis is proposed as a risk factor for preterm delivery, fetal growthrestriction and preeclampsia with severe consequences for maternal andneonatal health, however the biological mechanisms involved are elusive.Porphyromonas gingivalis (Pg), one of the main pathogens in periodontaldisease, produces extracellular vesicles referred to as outer membrane vesicles(PgOMV), which serve a relevant function in the interaction with host cells. In theprocess of placentation, trophoblast cells (Tb) release soluble factors whileclosely engaging with maternal leukocytes to maintain immune homeostasis.Disruption in this trophoblast-immune interplay has been linked to pregnancycomplications.Objective: The aim of this work is to study the effect of PgOMV on trophoblastimmune interaction.Methods: Trophoblast cells were exposed to PgOMV stained with PKH67fluorescent dye at different concentrations and time periods to examineinternalization. mRNA expression of inflammatory mediators was evaluated byqPCR. Neutrophils were purified from healthy donors and cultured withconditioned media from Tb (CM) pre-treated or not with PgOMV (PgOMV CM).Migration to CM was evaluated by flow cytometry using transwell assays andReactive Oxygen Species (ROS) production using DCFH-DA probe. Monocyteswere isolated from the peripheral blood of healthy female donors using FicollPaque/Percoll and were cultured with rhGM-CSF+rhIL-4 for 5 days in theabsence/presence of CM or PgOMV CM. Additionally, monocytes weredifferentiated into macrophages with rhM-CSF for 5 days. These macrophageswere then cultured for 48 hours with conditioned media from Tb CM or PgOMVCM. The macrophage and dendritic cell profile was assessed by flow cytometry.Results: PgOMV are internalized by trophoblast cells in a time and dosedependent manner. PgOMV treatment induces a selective modulation ofinflammatory mediators with reduced mRNA expression of IL-8, IL-6accompanied by an increased expression MCP-1 (p < 0.05). Trophoblast cellsprimed with PgOMV enhanced neutrophil chemoattraction (p < 0.05) andincreased ROS production (CM 15967 ± 2989; PgOMV CM 20068 ± 3982;X±SEM n=9; p < 0.05). Enhanced ROS production was associated with anincreased mRNA expression of gp91phox p47phox and p67phox, subunits of theNADPH oxidase complex and Glucose-6-phosphate dehydrogenase, the ratelimiting enzyme of the pentose phosphate pathway that fuels NADPH oxidasewith NADPH to produce superoxide (p < 0.05). PgOMV CM also affected theprofile of macrophages and dendritic cells.Conclusion: Taken together, results support a pathogenic role of PgOMV at earlystages of pregnancy and placentation through disrupting trophoblast contributionto immune homeostasis maintenance.