Lactobacilli inhibits the formation of neutrophil extracelullar traps in a PMA induction model.

HUGO, A. A.; GORGOJO, J. ; RODRÍGUEZ, M.E.; PÉREZ, P.F.

Mar del Plata

Congreso; LXVI Reunión Anual de la Sociedad Argentina de Inmunología; 2018

Sociedad Argentina de Inmunología

Neutrophils are part of the first line of immune defense. Neutrophils utilize different mechanisms to protect hosts such us phagocytosis, degranulation and production of neutrophil extracellular trap (NET). NETs are DNA web-like extracellular structures that immobilize and kill bacteria due to the adhesion of antimicrobial proteins. Uncontrolled NETosis leads to liberation of cell-free DNA and proteases that damage the surrounding cells and exacerbate inflammation. In the gut environment, the presence of beneficial microbes could reduce intestinal inflammation and protect the host. We investigated the capacity of two lactobacilli strains L. delbrueckii subsp lactis (CIDCA 133) and L. delbrueckii subsp bulgaricus (CIDCA 331) to inhibit NET formation.Human neutrophils were isolated from heparinized venous blood using gradient centrifugation. Cells were incubated with or without lactobacilli at different multiplicity of infection (moi). PMA was used as NET inducer. NETs were visualized by immunofluorescence microscopy. ROS was determined by luminometry. Results show that in the absence of stimuli lactobacilli lack the ability to induce NETs. Moreover, lactobacilli (MOI 10) have the ability to significantly reduce NETs formation in response to PMA challenge. Strain CIDCA 133 showed a stronger capacity to decrease the number of netotic cells than strain CIDCA 331 (66,09 % ± 3,09 vs 27,69 % ± 0,17, respectively) after PMA treatment. Lactobacilli showed different capacity to stimulate ROS generation, with strain CIDCA 133 as a strong inducer and strain CIDCA 331 as a non-inducer. Neither of these strains were able to inhibit ROS generation in PMA treated cells. Our results showed that lactobacilli strains had the ability to reduce NET formation but not to inhibit ROS generation in PMA treated cells. NET formation is tightly related to ROS generation and autophagy in PMA induced NETosis. It is possible that lactobacilli are affecting the autophagy process, therefore, reducing NET formation.