Presence Of Neutrophil Extracellular Traps In The Blood Of Bacterial Sepsis And Different Patterns Of NETs Generation Induced By Klebsiella Pneumonia In Vitro

Objective The aims of this study were to identify NETs in the blood of patients with sepsis and to investigate the responsiveness of human neutrophils to two different breeds of klebsiella pneumoniae focusing on NETs formation.Method A prospective study from May to November 2015 was performed on 60 adult patients who met criteria of sepsis to the definition accepted at the ACCP/SCCM conference,which including 30 patients with sepsis induced by k.pneumoniae.30 healthy individuals served as controls.Blood samples that obtained from 30 patients at day1 and 7 were stained with Sytox Green.So does with the healthy donors Circulating DNA and histone H3 were visualized simultaneously by immunofluorescence.In vitro,human neutrophils were challenged with hypermucoviscous Klebsiella pneumoniae(hv KP)and classic Klebsiella pneumoniae(c KP).Results 1.NETs were visualized with Sytox Green in the blood of sepsis patients and their individual components were evaluated simultaneously.2.NETs were abundant in the plasma of patients(n=30)on a day of recruitment compared to healthy controls(42.358%±1.967 vs 0.262%±0.041),(P<0.05);3.Patients which still suffering from severe infection had significantly higher NETs levels(51.191%±5.550)compared to the recovering patients(19.793%±1.917),(P<0.05);4.NETs from hv KP samples appeared to be bigger than those presented by c KP samples,which seemed to be smaller(53.865%±1.385vs27.628±1.425),(P<0.05);5.The formation of NETs was more prominent when neutrophils were challenged with hv KP than c KP(44.581%±2.225 vs 28.022%±1.416),(P<0.05);6.The survival rate of c KP was less than 40% in the presence of neutrophils.However,hv KP was above 70%(32.579%±1.155 vs 71.660%±1.446),(P<0.05).Conclusion This clinical study of blood samples report a number of advancements in understanding of NETs biology in host defense,including the demonstration the presence of NETs during bacterial sepsis,comparison of the quantification of NETs area on day of enrollment and day 7,and last but not least,investigation on neutrophils responsiveness to c KP and hv KP,focusing on NETs formation.Furthermore,our study expands on investigations of therapeutic targeting of NETs.