Induction of Apoptosis in Neutrophils from Septic Patients by Proinflammatory Molecules.

Neutrophils are in part responsible of the tissue damage accompanying the systemic inflammatory response produced during sepsis. A mechanism to limit their activity is apoptosis, however neutrophil apoptosis appear to be delayed in sepsis, which could increase the abnormal inflammatory response. On the other hand, it has been described that the proinflammatory molecules involved in delaying neutrophil apoptosis can induce apoptosis in other cells. Moreover there is not studies evaluating early apoptotic response in neutrophils from septic patients. In order to study the early apoptic response (30, 60 and 180 min) of neutrophils isolated from septic patients and healthy volunteers, these cells were isolated from peripheral blood, cultured with not stimulus or with LPS or fMLP and then analyzed by flow cytometry using DiOC6 and Annexin V/propidium iodide. Our data show that spontaneous apoptosis in neutrophils isolated from septic patients is increased during the first 3 h of in vitro culture in comparison to apoptosis from healthy volunteers neutrophils; however, this response is delayed when neutrophils isolated from septic patients were cultured during 24 h. Furthermore neutrophils isolated from healthy controls and septic patients stimulated with LPS or fMLP evidenced an apoptotic response at early incubation times in comparison with untreated cells. Previous studies have demonstrated a delay of apoptotic response of neutrophils obtained from patients with sepsis, which has been explained by the induction of anti-apoptotic proteins or inhibition of expression of apoptotic genes by LPS and other proinflammatory mediators. On the contrary our results suggest that there is an early increase in apoptosis when neutrophils isolated from septic patients are cultured without stimulus or in presence of microbial molecules such as fMLP and LPS. This contradictory evidence might be explained if we take in account the heterogeneity of circulatory neutrophils. At early time after LPS incubation a susceptible population of neutrophils activates different apoptotic responses, however the cells that are resistant to this effect activate a cascade of intracellular events that finally induces the transcription of several genes with anti-apoptotic and inhibits the expression of those pro-apoptotic. As consequence of these differences new pathogenic processes and therapeutic targets should be considered in sepsis.