Plasma Levels of IL-8 and Microparticle Tissue Factor Activity Are Associated with Mortality in Patients with Primary Influenza A/H1N1 Infection

Background and Objective:  Treatment and triage decisions during influenza remain difficult due to lack of reliable severity of illness predictive score. Influenza A/H1N1 induces the expression and release of tissue-factor bearing microparticles (MP-TF), contributing to a prothrombic milieu. However, there are no studies reporting levels of circulating TF-expressing MPs during the course of human influenza.  We sought to determine if MP-TF are an early predictor of mortality in critically ill patients with influenza A/H1N1.

Methods:  This was a prospective, multicenter, case-cohort pilot study of three academic intensive care units.  We prospectively studied 15 patients with primary influenza A/H1N1 that included 7 survivors and 8 non-survivors.  For comparison, 27 healthy, medication-free, age- and gender-matched control subjects were also prospectively studied.  Plasma was prepared from blood drawn upon ICU admission in influenza patients.  MP-TF activity, thrombin-antithrombin complexes (TATc), and D-dimers were measured as markers of activation of coagulation.  Plasma cytokine levels were measured on the same blood samples.  Patients were followed for the primary outcome of 28-day mortality.

Results:  The average admission APACHE II score of the influenza patients was 25.5±9.3, 60% of patients had shock, and the 28-day mortality rate was 53.3% (n=8/15).  Compared to healthy controls, influenza patients had significantly higher plasma fibrinogen, C-reactive protein (CRP) MP-TF activity, TATc, D-dimer and a prolonged prothrombin time.  However, of these procoagulant markers, only MP-TF activity predicted influenza related mortality (5.6±1.2 pg/ml in non-survivors vs. 1.8±0.8 pg/mL in survivors, p < 0.05; Table 1 and Fig. 1A).  MP-TF activity, TATc levels, and D-dimer did not correlate with APACHE II score, platelet count, fibrinogen levels, CRP, or age or between patients with severe sepsis versus septic shock.  Influenza non-survivors also had significantly higher plasma IL-8 levels compared with survivors (71.8±29.1 pg/ml vs. 17.3±3.7 pg/mL, p < 0.05; Figure 1B).  MP-TF activity and IL-8 levels were significantly and positively correlated (r² = 0.60, P=0.003; Figure 1C).  Other cytokines, TATc, and D-dimer were not different between non-survivors and survivors.

Conclusions:  This study demonstrates that plasma IL-8 and MP-TF activity measured upon admission in patients with severe, primary influenza A/H1N1 infection is associated with subsequent mortality.  Thus, these biomarkers may serve as very early prognostic markers for patients with influenza A/H1N1.

 Table 1.  Characteristics of the influenza patients.  Laboratory values, coagulation markers, and plasma cytokines were measured within 24 hours of ICU admission.

	All Influenza A/H1N1 Patients (n=15)	Influenza Non-Survivors (n=8)	Influenza Survivors (n=7)	P value
Admission Characteristic
Age, years	43.3±11.0	45.8±4.6	40.4±15.5	0.37
Male Gender, n (%)	7 (47%)	3 (38%)	4 (57%)	0.45
Weight, kg	95.5±25.2	86.6±20.9	105.7±27.4	0.15
BMI, kg/m2	33.3±7.2	31.5±5.0	35.4±9.1	0.31
Obesity (BMI³30 kg/m2)	10 (67%)	5 (63%)	5 (71%)	0.71
Tobacco Use, n (%)	4 (27%)	3 (38%)	1 (14%)	0.31
APACHE II Score	25.5±9.3	27.9±9.1	22.9±9.4	0.31
Mechanically Ventilated, n (%)	15 (100%)	8 (100%)	7 (100%)	--
Shock, n (%)	9 (60%)	6 (75%)	3 (43%)	0.21
P/F Ratio	83±28	84±34	82±22	0.89
Clinical Outcomes
ICU Length of Stay, days	21.9±7.7	23.0±7.9	20.7±7.8	0.58
Duration of Ventilation, days	8.2±1.0	8.5±0.9	7.6±0.9	0.11
Secondary Bacterial Infection, n (%)	5 (33%)	4 (50%)	1 (14%)	0.28
Overt DIC, n (%)	15 (100%)	8 (100%)	7 (100%)	--
Laboratory Values
Platelets, 103/µL	213±138	154±104	280±150	0.08
White Blood Cells, K/µL	7.3±4.4	8.5±4.9	5.9±3.5	0.28
Hemoglobin, g/dL	11.7±1.9	11.5±2.0	12.0±1.8	0.66
Serum Creatinine, mg/dL	1.1±0.7	1.14±0.74	1.07±0.59	0.84
Coagulation Markers
Fibrinogen, mg/dL	571±240	461±253	700±157	0.07
C-reactive Protein, mg/L	11.0±7.6	11.8±7.4	10.2±8.2	0.70
PT, sec	20±7.0	19.2±4.7	20.9±9.3	0.67
aPTT, sec	53.0±22.9	55.1±29.0	50.7±15.0	0.72
MP-TF, pg/mL	3.8±0.9	5.6±1.2	1.8±0.8	< 0.05
TATc, ng/mL	11.8±2.7	14.3±3.5	9.1±4.1	0.35
D-dimer, ng/mL	2439±86	2568±98	2292±134	0.11

Figure 1. MP-TF activity and IL-8 predict mortality in patients with influenza A/H1N1. 

(A) Plasma levels of MP-TF activity and (B) IL-8 levels in survivors and non-survivors. (C) Correlation between MP TF activity and IL-8 levels in H1N1 influenza-infected patients (*P<0.05).