IntroductionSubstantial units of fresh frozen plasma (FFP) are utilized in the intensive care unit (ICU) [1,2]. FFP is effective in correcting clotting factor deficiencies [3] and is therefore transfused in patients with active bleeding, but also frequently in patients with abnormal coagulation tests to prevent bleeding [2,4]. In sepsis patients, FFP transfusion rates of up to 57% have been reported [5]. However, there is an association between FFP transfusion and adverse outcome in the critically ill, including transfusion-related acute lung injury (TRALI) [4,6-8], transfusion-related circulatory overload [9,10], multiorgan failure [8,11] and an increased risk of infections [12]. Although not entirely understood, the pathological mechanisms underlying the association between FFP transfusion and lung injury is thought to result from an inflammatory response including a neutrophil influx into the lungs and elevated pulmonary levels of interleukin 8 (IL-8) and interleukin 1 (IL-1), as demonstrated in TRALI patients [13,14]. In line with this, FFP increased expression of endothelial adhesion molecules in human pulmonary endothelial cells [15]. Together, these data suggest that endothelial cell activation and disruption may be an early event following lung injury due to transfusion [16].On the other hand, FFP also seems to have protective effects. In trauma patients requiring a massive transfusion, resuscitation with a higher ratio of FFP to red blood cell units is associated with decreased mortality [17,18]. Interestingly, some studies suggest that this decreased mortality is irrespective of correction of coagulopathy by restoring coagulation factor levels [18,19], although not all studies support this observation [20,21]. Instead, a beneficial effect of FFP may be related to the restoration of injured endothelium. Syndecan-1 is a proteoglycan on the luminal surface of endothelial cells that inhibits leukocyte adhesion. During endothelial damage, syndecan-1 is shed, resulting in increased levels of syndecan-1 in the systemic compartment [22]. Patients in hemorrhagic shock have a disrupted endothelial integrity and glycocalyx layer, with decreased syndecan-1 expression [23]. Vascular integrity is also disrupted in various populations of critically ill patients, as demonstrated by increased systemic levels of syndecan-1 [24,25]. Of interest, in a hemorrhagic shock model, FFP was found to improve endothelial integrity, associated with increased expression of syndecan-1 on endothelial cells [26].The effect of transfusion of FFP on endothelial and cytokine host response in patients is unknown. In a study investigating the risk-benefit ratio of FFP transfusion in non-bleeding critically ill patients with a coagulopathy, we investigated the inflammatory and endothelial host response to a fixed dose of FFP transfusion.MethodsStudy designThis was a predefined post hoc substudy of a multicenter trial in which non-bleeding critically ill patients with an increased international normalized ratio (INR, 1.5 to 3.0) were randomized between May 2010 and June 2013 to omitting or administering a prophylactic transfusion of FFP (12 ml/kg) prior to an invasive procedure. Only patients randomized to receive FFP were included in this substudy. Patients were enrolled at three sites in The Netherlands: two university hospitals (Academic Medical Center, Amsterdam and Leiden University Medical Center, Leiden) and one large teaching hospital (Tergooi Ziekenhuizen, Hilversum). The Institutional Review Board of the Academic Medical Center approved the study protocol. Before entry in the study, written informed consent was obtained from the patient or legal representative in accordance with the Declaration of Helsinki. The study protocol was registered with trial identification numbers NTR2262 and NCT01143909 [27].Exclusion criteria were clinically overt bleeding, thrombocytopenia of <30 × 109/L, treatment with vitamin K antagonists, activated protein C, abciximab, tirofiban, ticlopidine or prothrombin complex concentrates and a history of congenital or acquired coagulation factor deficiency or bleeding diathesis. Patients treated with low-molecular-weight heparin (LMWH) or heparin in therapeutic dose were eligible if medication was discontinued for an appropriate period. Sepsis was defined by the Bone criteria [28]. Disseminated intravascular coagulation (DIC) was defined by an International Society on Thrombosis and Haemostasis (ISTH) score of ≥5 [29]. The FFP was quarantine plasma manufactured by Sanquin, the Dutch National Blood Bank. As of 2007, women are deferred for donation for preparation of FFP in the Netherlands.Sample collectionCitrated blood samples were drawn from an indwelling arterial catheter before and within 10 minutes after FFP transfusion. During transfusion, respiratory settings were kept constant. Samples were collected in sodium citrate (0.109 M 3.2%) tubes and were centrifuged twice within 30 minutes: the first 15 minutes at 2,000 × g and then 5 minutes at 15,000 × g, both at 18°C. Supernatant was collected and stored at −80°C.AssaysTumor necrosis factor alpha (TNF-α) levels were measured by enzyme-linked immunosorbent assay (ELISA), according to instructions of the manufacturer (R&D Systems Inc., Minneapolis, MN, USA). Serum levels of interleukin 1 beta (IL-1β), interleukin 1 receptor antagonist (IL-1RA), IL-8, interleukin 10 (IL-10), macrophage inflammatory proteins (MIP)-1A, monocyte chemotactic protein (MCP)-1 and soluble CD40 ligand (sCD40L) were determined by Luminex, according to the manufacturer’s instructions (Merck Millipore Chemicals BV, Amsterdam, The Netherlands). When less than 50 beads were measured by the Luminex assay, samples were excluded from further analysis. von Willebrand factor antigen (vWF:Ag) levels were determined with an in-house ELISA using commercially available polyclonal antibodies against von Willebrand factor (vWF) (Dako, Glostrup, Denmark). ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) activity was determined as described earlier [30]. Factor VIII activity was determined on a Behring XP coagulation analyzer using reagents and protocols from the manufacturer (Siemens Healthcare Diagnostics, Marburg, Germany).Statistical analysisVariables are expressed as medians and interquartile ranges (IQRs) or means and standard deviations (SDs). For comparisons, a paired t test was used, or the Wilcoxon signed-rank test in case of not normally distributed data. For the analyses, we used SPSS version 21.0 (IBM Corp., Armonk, NY, USA) and Graphpad Prism 5 (Graphpad Software Inc., San Diego, CA, USA).ResultsPatientsFrom 38 patients receiving FFP, paired samples from 33 patients were available for analysis before and after FFP transfusion. Patients were ill, as reflected by a high disease severity score and half of the patients had sepsis (Table 1). Patients received a mean dosage of 11.2 (2.8) ml/kg FFP, which was transfused in 121 ± 43 minutes.Table 1 , Fresh frozen plasma (FFP) is indicated for the management of massive bleedings. Recent audits suggest physician knowledge of FFP is inadequate and half of the FFP transfused in critical care is inappropriate. Trauma is among the largest consumers of FFP. Current trauma resuscitation guidelines recommend FFP to correct coagulopathy only after diagnosed by laboratory tests, often when overt , Introduction Much controversy exists on the effect of a fresh frozen plasma (FFP) transfusion on systemic inflammation and endothelial damage. Adverse effects of FFP have been well described, including acute lung injury. However, it is also suggested that a higher amount of FFP decreases mortality in trauma patients requiring a massive transfusion. Furthermore, FFP has an endothelial .