Avaliação prospectiva das atividades fagocitária e quimiotática de neutrófilos humanos quando submetidos ao plasma de pacientes sépticos

Abstract

Innate immune responses are altered during sepsis and the influence of septic plasma components over phagocytosis and chemotaxis remains to be determined. We sought to investigate the effects of the plasma from patients with sepsis over the phagocytic and chemotactic activities of neutrophil-like HL-60 cells, comparing these results with those observed with plasma from patients with noninfectious systemicinflammatory response (SIRS) and patients without SIRS. Plasma was obtained at the time of sepsis or SIRS diagnosis (day 1), and on days 2, 3, 5, 10, 20 and 28. Phagocytosis tests were performed by adding plasma to neutrophil-like HL-60 differentiated cells and fluorescent E.coli. The phagocytic index was measured by flow cytometry after 20 and 50 incubation. The chemotaxis tests were performed in response to fMLP and IL-8 by incubating plasma and differenciated HL-60 cells. Migrating cells were counted by using fluorometric microvolume assay technology. 66 patients were included, sepsis (n=30), non-infectious SIRS (n=24), and non-SIRS (n=11). The phagocytic activity of neutrophils at 20 and 50 measured at inclusion was significantly higher with plasma obtained from septic patients as compared to that observed when plasma from noninfectious SIRS and non-SIRS patients were used (p=0.009, p=0.03). This difference remained significant after 50 with the plasma obtained on days 2 and 5 (p=0.02, p=0.001, respectively). Considering only patients with SIRS (n=54), the phagocytic activity was significantly higher after 20 min in the septic group with plasma obtained at inclusion (p=0.01), with a trend to significance after 50 incubation (p=0.06). In the 12 septic patients alive at day 28, the phagocytosis activity increased during the follow up, and the difference between the phagocytic indexes measured at inclusion and on 28th day was significant for both incubation times (p=0.04 for both). The chemotactic activity of neutrophils in response to fMLP and IL-8 measured at inclusion was significantly decreased by septic plasma in comparison with non-infectious SIRS and non-SIRS groups (p=0.007, p<0.0001, respectively). Plasma from patients with sepsis decreased IL-8-dependent chemotactic activity more than that obtained in patients with SIRS, at the time of inclusion, on day 2, and on day 3 (p=0.05, p=006, p=0.005, respectively). In septic patients alive on day 28, the chemotactic response to IL-8 increased over the time, and the difference between the time of inclusion and the 28th day was significant (p=0.001). Remarkably, for all collecting points, there was no difference in IL-8 plasma levels between septic and non-infectious SIRS groups. Higher levels of bacterial phagocytosis and lower levels of chemotaxis to IL-8 were associated with increased 28-day mortality. Taking in account all studied patients, a significant correlation between phagocytic / chemotactic activities and severity (SAPS III and SOFA scores) was observed. All told, we can conclude that plasma from septic patientsenhances the phagocytic activity and impairs the chemotaxis of neutrophil-like cells. The phagocytic function remains increasing during the follow-up of septic survivors while chemotactic activity progressively recovers along sepsis evolution, reaching supra normal leels at the end of follow-up. Opsonins and mediators present in septic plasma influencing innate response remain to be unraveled.