Methods
intraoperative bleeding, blood loss, neoplasia, infection, hemolysis
BLOOD LOSS, NEOPLASIA, CHRONIC INFLAMMATORY DISEASE, INFECTION, BLOOD, HEMOLYSIS, INTRAOPERATIVE BLEEDING
This single center pilot study (NCT02674906) was performed in the operating room and the mixed medical-surgical ICU of Ziekenhuis Oost Limburg (ZOL), Genk, Belgium, a 805-bed non-university teaching hospital. This study was approved by the local ethics committee and written informed consent was obtained prior to surgery from the patient or legal representative. After consent, patients were randomized preoperatively either to a citrate anticoagulant group or a heparin anticoagulant group for cell salvage. During CABG, the perfusionist and anaesthesiologist were unblinded for the anticoagulation regimen used. Blinding was complete for the patient, as well as personnel in the intensive care unit (ICU) and ward.Adult patients undergoing elective on-pump CABG were considered eligible for inclusion. Elective surgery was defined as planned at least 24 h before surgery. Off-pump CABG, urgent procedures, use of vasoactive medication prior to surgery, infection treated with antimicrobial therapy, chronic inflammatory disease, immune suppressive drug treatment, active neoplasia, renal replacement therapy, or use of extracorporeal membrane oxygenation (ECMO) were exclusion criteria. In addition, we excluded patients who had massive intraoperative bleeding that could not be safely managed while collecting study data. Patients could be excluded at any time if the inclusion criteria were no longer met or when exclusion criteria appeared. Patients were randomized using randomly generated treatment allocations within sealed opaque envelopes.CS is a procedure to collect blood lost in the surgical field for reuse. Blood is collected in cell saver blood collection reservoir (BCR), where it is mixed with an anticoagulant, either citrate or heparinized saline. The collected blood-anticoagulant mixture is processed by filtering, after which it is drawn into a centrifuge. Isotonic saline solution is added to the centrifuge bowl as washing fluid. The centrifugal procedure separates red blood cells, which are denser and are propelled against the outer wall of the bowl, while less dense plasma moves towards the centre of the bowl where it is deposited in a waste bag. Waste products, including white blood cells, platelets, plasma, anticoagulant, fat, clotting factors, and free plasma haemoglobin are collected in the waste fluid. The washed red blood cell concentrate (WRBC) is collected in a separate bag.Anaesthesia, cardiopulmonary bypass and cell salvage procedures were all executed according to the standard protocol used in our institution (see Additional file Washed red blood cell concentrate (WRBC) was transfused per-operatively or immediately post-operatively. If there was ongoing blood loss, the standard hospital protocol was applied to manage this; if there was necessity for allogenic blood transfusion, this was administered irrespective of the type of anticoagulant used (the ICU ward and the intensivist were blinded to the anticoagulant group).The standard protocol left transfusion at the discretion of the board certified anesthesiologist and/or ICU staff member; a Hb < 8 g/dl in the peripheral blood is in our protocol considered the threshold for allogenic blood transfusion in this type of patients.We collected baseline characteristics of patients before CABG. We measured effects of the specific anticoagulation protocol on quality of WRBC, and at various time points inflammatory parameters and variables of hemolysis and coagulation.On one hand, the quality of WRBC was analysed by measuring hemoglobin (Hb), hematocrit (Ht), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), red cell distribution width (RDW), free hemoglobin (fHb), iron, thrombocytes, white blood cell (WBC) count and differentiation in WRBC compared to Hb and Hct in the BCR. Inflammatory parameters were analysed by measuring levels of interleukin (IL)-6, IL-8, IL-10 and myeloperoxidase (MPO) at baseline, in the BCR (in the blood-anti-coagulant mixture before processing) and in peripheral blood of the patient (taken from the arterial catheter in place) at two time points; Firstly, when the patient was successfully weaned from cardiopulmonary bypass, protamine had been given and when the WRBC had not yet been transfused. Secondly, in the ICU, 2–3 h post transfusion of WRBC, but before extubation or allogenic blood transfusion.The impact after transfusion of WRBC was evaluated, apart from measuring inflammatory parameters, by measuring fHB, iron, transferrin, ferritin, haptoglobin, hepcidin, prothrombin time (PT), activated partial thromboplastin time (aPTT), international normalized ratio (INR), rotational thromboelastometry (ROTEM) in peripheral blood of the patient (taken from the arterial catheter), at the same two time points before and after transfusion of WRBC. Furthermore, blood loss in thoracic drains and transfusion requirements over the first 24 postoperative hours were compared between groups.For determining the levels of interleukins and MPO, validated and commercially available assays were used: IL-6: Electrochemiluminescence immunoassay, (Elecsys Il-6 on Cobas e801); IL-8: human IL-8 ELISA kit (cat.no. KHC0081), Thermo Fischer Scientific; IL-10: human IL-10 ELISA kit (cat. No BMS215-2) Thermo Fischer Scientific; MPO: Myeloperoxidase ELISA kit (ref: KT-890) Epitope Diagnostics, inc. All assays were performed according to the manufacturers’ instructions, with internal quality controls and participation to external quality control programs.Statistics were performed using SPSSA sample size of 34 achieves 80% power to detect a difference of 2 g/dl in Hb, a primary outcome parameter, using a two-tailed two sample t-test at a 0.05 level of significance. Equal sized groups and a standard deviation of 2 was assumed.