OC 75.3 - Fibrinogen binds circulating histones in vivo in the crosstalk between coagulation and innate immunity.

Background: Fibrinogen binds extracellular histones in vitro.

Aims: It is unclear how this translates in vivo and affect patient outcomes.

Methods: C57/BL6 mice (6-8/group) were infused with fibrinogen (100mg/kg) or saline (control) before receiving histones (75mg/kg). For organ injury assessments, blood and organs were harvested 2-6 hours post-infusion. For survival analysis, mice were observed for 7 days post-infusion.

An ELISA method was used to detect circulating histone-fibrinogen complexes. 96-well plates were coated with anti-histone single-chain fragment variable (ahscFv) and incubated with human plasma, before being probed with an anti-human fibrinogen antibody.

To determine clinical significance, an admission ratio of histone (µg/mL)-to-fibrinogen (g/L) (histone:fibrinogen) was calculated in a cohort of critically ill patients. Its predictive value was then assessed by binary regression modelling using a previously established cut-off of 6.

Results: In mice, fibrinogen pre-infusion significantly reduces histone-induced thrombocytopenia and organ injury compared to saline controls (median±SD; platelet count 333.80±95.99x109/L vs 847.57±115.93x109/L, p< 0.001; cardiac troponin I 0.06±0.05ng/mL vs 0.02±0.01ng/mL, p=0.038; creatinine 26.80±10.03ng/mL vs 10.55±6.04ng/mL, p=0.014; alanine transaminase 650.20±243.15ng/mL vs 177.55±67.28ng/mL, p=0.011; and lung injury score 0.16±0.06 vs 0.34±0.1, p=0.039). All mice receiving histone infusion alone died within 24 hours while all mice receiving fibrinogen pre-infusion survived.

Circulating histone-fibrinogen complexes were detectable and significantly (p < 0.001) elevated in most critically ill patients (n=75, optical density [OD]=0.214 [0.178-0.257]) when compared to normal controls (n=18, OD=0.155 [0.134-0.172]). An admission histone:fibrinogen ≥6 predicted development of moderate-severe thrombocytopenia (platelet count < 100x109/L). This was also predictive of 28-day mortality by univariate analysis [odds ratio=2.267 (95% CI 1.119-4.590, P=0.023)] and multivariate analysis following adjustment for age, gender and ethnicity [odds ratio=2.191 (95% CI 1.071-4.485, P=0.032)].

Conclusion(s): This study advances the field by translating in vitro observations of fibrinogen-histone binding into in vivo relevance. Furthermore it has clinical relevance as histone:fibrinogen levels on ICU admission has prognostic significance.