Effects of short-term fever-range hyperthermia on neutrophil response to Pseudomonas aeruginosa

KEITELMAN, IRENE; SABBIONE, FLORENCIA; GIAI, CONSTANZA; TOWSTYKA, NADIA YASMÍN; GUZMÁN, MAURICIO; GALLETTI, JEREMÍAS; JANCIC, CAROLINA CRISTINA; GÓMEZ, MARISA; TREVANI, ANALÍA S.

Mar del Plata

Congreso; 64 Reunión Anual de la Sociedad Argentina de Inmunología; 2016

Fever response is a hallmark of infection and inflammatory disease, working as an alert system that promotes immune surveillance during challenge by invading pathogens. Previously we determined that 1h of hyperthermic conditions (39.5°C) significantly increased NETosis induced by PMA. However, cytokine release in response to LPS was significantly inhibited. In this study, we aim to determine if short term hyperthermia is able to modulate NET release and other neutrophil (PMN) functions in response to the opportunist pathogen Pseudomonas aeruginosa (Pa). HumanPMN were incubated with or without Pa PAO-1 strain (MOI: 1:1) at 37°C or 39.5°C for 1h, and then cultured for different periods at 37°C according to the evaluated function. The secretion of IL-8 was determined by ELISA and elastase secretion was assayed by degradation of a specific substrate. NETosis was evaluated y determining co-localization of elastase and DNA by confocal microscopy (CM), and DNA released by fluorometry. Autophagy levels were determined by quantification of vesicular LC3B/cell by CM. Results indicated that 1h of exposure to 39.5°C and additional culture at 37°C did not modulate the capacity of bacteria to induce DNA release at 1, 2, 3 or 4h (n=4). Hyperthermia did not affect NET release induced by Pa evaluated by CM at 2, 3 and 4h post-thermic treatment (n=2). However, elastase levels detected in Pa-stimulated-PMN supernatants were reduced in cells exposed to hyperthermia (n=4; p<0.05) suggesting that part of it could have been released not only by NETosis but also during bacterial phagocytosis. Similar findings were obtained when autophagy levels were evaluated (n=4). As it was observed using LPS as stimulus, hyperthermia inhibited IL-8 secretion induced by Pa evaluated at 2, 3, or 4h post-stimulation (n=3). In summary, these results suggest that short-term hyperthermia differentially affects not only NETosis in response to PMA and Pa, but also distinct neutrophils responses induced by Pa.