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Clinical Science  |   November 1999
Perflubron Emulsion Delays Blood Transfusions in Orthopedic Surgery 
Author Affiliations & Notes
  • Donat R. Spahn, M.D.
    *
  • Ronald van Brempt, M.D.
  • Gregor Theilmeier, M.D.
  • Jan-Peter Reibold, M.D.
    §
  • Martin Welte, M.D.
  • Hartmut Heinzerling, M.D.
    #
  • Katrin M. Birck, M.D.
    **
  • Peter E. Keipert, Ph.D.
    ††
  • Konrad Messmer, M.D.
    ‡‡
  • *Professor, Department of Anesthesiology, University Hospital Zürich, Zürich, Switzerland. †Assistant Professor, Department of Anesthesiology, University Hospital Leuven, Leuven, Belgium. ‡Assistant Professor, Department of Anesthesiology, University Hospital Münster, Münster, Germany. §Assistant Professor, Department of Anesthesiology, University Hospital Giessen, Giessen, Germany. ∥Associate Professor, Department of Anesthesiology, University Hospital Munich, Munich, Germany. #Clinical Research Manager, R. W. Johnson Pharmaceutical Research Institute, Zürich, Switzerland. **Director of Clinical Research, R. W. Johnson Pharmaceutical Research Institute, Zürich, Switzerland. ††Program Director, Oxygen Carriers Development, Alliance Pharmaceutical Corporation, San Diego, California. ‡‡Professor for Experimental Surgery, and Director, Institute for Surgical Research, University of Munich, Munich, Germany. §§Members of the European Perflubron Emulsion Study Group are listed in Appendix 1.
Article Information
Clinical Science
Clinical Science   |   November 1999
Perflubron Emulsion Delays Blood Transfusions in Orthopedic Surgery 
Anesthesiology 11 1999, Vol.91, 1195. doi:
Anesthesiology 11 1999, Vol.91, 1195. doi:
THE risks and side effects of allogeneic blood transfusions include transfusion reactions, 1 alloimmunization, 2 transmission of infectious agents 3,4 and immunosuppression 5 resulting in an increased incidence of postoperative infections 6 with prolonged hospitalization, 7 higher costs, 8 and probably a shorter period of recurrence-free survival after cancer surgery. 9 Therefore, to avoid unnecessary blood transfusions, the use of physiologic transfusion triggers rather than exclusively hemoglobin-based transfusion triggers has been suggested. 10–12 
Both preoperative autologous blood donation and acute normovolemic hemodilution (ANH) have been proposed to minimize allogeneic blood transfusion requirements. The efficiency of each method, however, has been challenged. The cost effectiveness of preoperative autologous blood donation has been questioned, 13 but it is the blood-sparing efficiency of ANH that has been debated. 14 ANH exposes the patient to lower hemoglobin concentrations, which may represent a risk for certain groups of patients. Therefore, combining ANH with an artificial oxygen carrier appears to be particularly intriguing: ANH could be extended to lower hemoglobin concentrations with an increased blood-saving efficiency, 15 and simultaneously improved safety during more profound anemia might be provided by the extra oxygen-delivery capacity of the artificial oxygen carrier if there are no adverse consequences of its use. 16,17 
Perfluorochemicals represent one group of artificial oxygen carriers intensively studied in recent years. 18 A second-generation concentrated perfluorochemical emulsion formulation undergoing advanced clinical testing is perflubron emulsion, a lecithin-based emulsion of perfluorooctyl bromide (C8F17Br) and perfluorodecyl bromide (C10F21Br) in a continuous aqueous phase without colloid consisting of phosphate buffered saline. Perflubron emulsion is characterized by a high dissolving capacity for gases such as oxygen and carbon dioxide, with a linear relationship between oxygen partial pressure and oxygen content, in contrast to the sigmoidal oxygen dissociation curve of hemoglobin. 16,19 Pharmacokinetic studies in human volunteers have yielded a blood half life of 9.4 ± 2.2 h (at a dose of 1.8 g/kg). 20 The purpose of this study was to evaluate safety and efficacy of perflubron emulsion in the setting of ANH to reverse physiologic transfusion triggers in patients undergoing orthopedic surgery with significant blood loss.
Materials and Methods 
Study Population 
The study was performed in 11 centers in six European countries in 147 of 290 formally screened patients reaching randomization between January 1996 and February 1997. The protocol was approved by all local ethics committees. Patients of either sex undergoing unilateral (first-time or revision) hip replacement or first-time spinal surgery with an expected blood transfusion requirement of 2–6 units of packed red cells were eligible for this study.
Additional inclusion criteria were a preoperative hemoglobin concentration between 11 and 16 g/dl; use of general, balanced anesthesia (isoflurane in oxygen and air with fentanyl and vecuronium) as opposed to spinal or epidural anesthesia; consent for insertion of a pulmonary artery catheter; acute normovolemic hemodilution (ANH) to a target hemoglobin concentration of 9 g/dl; an arterial oxygen partial pressure ≥ 360 mmHg after a 5-min ventilatory trial at an inspiratory fraction of oxygen (FIO2) of 1.0; and the ability to achieve a mixed venous oxygen partial pressure (PvO2) of ≥ 40 mmHg after ANH at an FIO2of 0.40. This PvO2was measured between the end of ANH and the beginning of surgery and was not always identical with the PvO2measured immediately after ANH.
Exclusion criteria were pregnancy or lactation; refusal of allogeneic blood transfusions; erythropoietin therapy within 21 days before surgery; hypersensitivity to constituents of the perflubron emulsion (e.g.  , egg yolk allergy); surgical procedure expected to last longer than 8 h; expected transfusion requirements greater than 6 units; spinal or epidural anesthesia (alone or as adjunct to general anesthesia); preoperative blood transfusions; congestive heart failure greater than New York Heart Association class II; baseline electrocardiogram with nonassessable ST segments (left bundle branch block, duration of QRS complex > 0.12 s, preexisting ST-segment depression); severe chronic obstructive pulmonary disease necessitating regular use of bronchodilators or corticosteroids; American Society of Anesthesiologists class > 3; platelet count < 150,000/μl; significant hepatic disease (aspartate aminotransferase or alanine aminotransferase > twice upper limit of normal); significant impairment of renal function (creatinine concentration > 180 μM); body weight > 175% of that recommended by Metropolitan Life Height and Weight tables for patient's height; use of preoperative oral anticoagulation; treatment with aspirin, fibrinolytic drugs, or antifibrinolytics; history of bleeding disorder; immunosuppression; local or systemic infection; history of malignancy within past 5 yr; and alcohol or drug abuse within the past year. Preoperative autologous blood donation was allowed, as well as fractionated low molecular weight heparin for the prophylaxis of postoperative deep vein thrombosis. Emulsion-based drugs other than perflubron emulsion were not administered in the perioperative period.
Study Protocol 
This was a prospective, multinational, multicenter, randomized, controlled, single-blind, parallel group study. Perflubron emulsion (Oxygent, Alliance Pharmaceutical, San Diego, CA) was provided by the manufacturer, and the study was sponsored by the R. W. Johnson Pharmaceutical Research Institute (Zürich, Switzerland, and Raritan, New Jersey). Perflubron emulsion has an oxygen solubility of 7.6 mM (19.3 vol%) and a carbon dioxide solubility of 61.6 mM (157 vol%) at 37°C and one atmosphere ambient pressure.
A peripheral venous cannula was inserted and anesthesia was induced. The anesthesia induction agent was not specified by the protocol. Anesthesia was maintained with isoflurane and fentanyl and patients were paralyzed with vecuronium. Patients were ventilated with oxygen-enriched air without nitrous oxide. No epidural or spinal anesthesia was allowed, either as a primary anesthesia or as an adjunct to general anesthesia.
Patients were monitored with radial artery and pulmonary artery catheters. After anesthesia induction, ANH to a target hemoglobin concentration of 9 g/dl was performed by blood withdrawal and simultaneous colloid replacement (gelatin, hydroxyethyl starch 200 kDa/0.5 M, or human serum albumin). After ANH a first set of hemodynamic measurements was performed at an FIO2of 0.40, including heart rate, systemic and pulmonary artery pressures, pulmonary capillary wedge pressure (PCWP), cardiac output (thermodilution), arterial and mixed venous blood-gas analyses, and hemoglobin concentration (HemoCue, AB Leo Diagnostics, Helsinborg, Sweden). The individual PCWP after ANH, measured in the position in which the operation was performed, served as an estimate of normovolemia in a given patient. During the subsequent operation, by protocol, PCWP was required to be kept at least at this posthemodilution level with crystalloid and colloid infusions. FIO2was maintained at 0.4 until randomization (fig. 1).
Fig. 1. Study protocol. 
Fig. 1. Study protocol. 
Fig. 1. Study protocol. 
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Most of the transfusion triggers were based on post-ANH hemodynamic measurements and are termed physiologic transfusion triggers  here. These physiologic transfusion triggers were prospectively defined in the protocol as tachycardia (heart rate > 125% of post-ANH heart rate or > 110 beats/min, whichever came first), hypotension (mean arterial pressure < 75% of post-ANH mean arterial pressure or ≤ 60 mmHg), high cardiac output (>150% of post-ANH cardiac output) or low PvO2(<38 mmHg). A hemoglobin concentration < 6 g/dl before reaching any of these physiologic transfusion triggers or electrocardiographic signs of myocardial ischemia (new ST-segment depression ≥ 0.1 mV or new ST-segment elevations ≥ 0.2 mV) prevented randomization and mandated retransfusion of autologous blood collected during ANH. If the patient had been randomized and one of the treatments had been given, a hemoglobin concentration < 6 g/dl or electrocardiographic signs of myocardial ischemia were considered additional triggers for subsequent blood transfusions.
Investigators were instructed to accept tachycardia and hypotension as physiologic transfusion triggers only after hypovolemia was excluded, as evidenced by a PCWP at or above the individual post-ANH PCWP level, and if an adequate depth of anesthesia was assured. This meant that increasing heart rates were to be treated first by fentanyl, and decreasing blood pressure by lowering isoflurane administration.
During surgery, the hemodynamic and blood-gas measurements were repeated at 30-min intervals or performed if a transfusion trigger was detected by continuous monitoring. After a transfusion trigger was reached, patients were randomized into one of four treatment groups based on a computer-generated randomization schedule using permuted blocks and stratification by center. Treatment groups were (1) a standard-of-care group, in which 450 ml of autologous blood (last complete unit removed during ANH) was transfused at an FIO2of 0.40 (the AB group);(2) a low-dose perflubron emulsion group, in which 0.9 g perfluorochemical per kilogram (1.5 ml/kg) and colloid added to a total volume of 450 ml were administered in conjunction with changing FIO2to 1.0 (the P0.9group);(3) a high-dose perflubron emulsion group, in which 1.8 g perfluorochemical per kilogram (3.0 ml/kg) and colloid added to a total volume of 450 ml were administered in conjunction with changing FIO2to 1.0 (the P1.8group); and (4) a colloid group, in which 450 ml of colloid was infused in conjunction with changing FIO2to 1.0 (the COL group)(fig. 1).
After completion of treatment and every 15 min thereafter, all previously mentioned hemodynamic and blood-gas measurements, including samples for perflubron concentration in the perflubron emulsion groups, were performed until a second transfusion trigger was reached. Upon reaching the second transfusion trigger all patients received a unit (450 ml) of autologous blood harvested during ANH (if available) or one unit of packed red cells (autologous or allogeneic) with ventilation at an unchanged FIO2according to treatment-group assignment. Cell salvage was allowed during the operation. The protocol mandated retransfusion of autologous blood, in the order of blood collected during ANH, cell saver blood, and blood from preoperative autologous donation, before using allogeneic blood. After the second treatment, hemodynamic and blood-gas measurements, including perflubron concentration in the perflubron emulsion groups, were performed at 60-min intervals or upon reaching the next transfusion trigger when autologous blood collected during ANH or one unit of packed red blood cells (autologous or allogeneic) was given. After initiation of wound closure, the hemoglobin concentration was adjusted to ≥ 8 g/dl by gradual transfusion of all remaining autologous blood before allogeneic blood had to be used.
Patients underwent follow-up evaluation from enrollment to postoperative day 28. Besides vital signs and general aspects of well-being, blood was collected 4 h after the first treatment and at postoperative days 1, 2, 3, 4, 7, 14 and 28 for the determination of laboratory profiles (hematology, blood coagulation, and chemistry). All adverse and serious adverse events were recorded during the first 28 postoperative days. Treatment-emergent events were defined as those that were new in onset (e.g.  , new onset of hypertension) or aggravated in frequency or severity following dosing (e.g.  , known hypertension requiring more intense treatment). In addition, any pathologic finding that was new in occurrence or exacerbated compared with the subject's status at study entry was considered a treatment-emergent adverse event if it required medical intervention.
The primary endpoint was the duration of reversal of changes in protocol-specified transfusion triggers, that is, the time from the start of the first treatment to the start of the second treatment or the end of surgery (initiation of wound closure), whichever occurred sooner. Secondary endpoints included percentage of patients achieving complete reversal of physiologic transfusion triggers; the need for allogeneic blood transfusions; oxygen dynamics including calculated oxygen content, oxygen consumption, oxygen delivery, and oxygen extraction (see Appendix 2), assessment of a dose response (low vs.  high perflubron dose); and safety evaluation in the postoperative period until postoperative day 28 including vital signs, laboratory profiles (hematology, blood coagulation, and chemistry), and adverse event data.
Statistics 
All analyses were performed on an intent-to-treat basis. The primary efficacy variable was duration of transfusion-trigger reversal. Subjects who did not reach a second transfusion trigger before the end of surgery were treated as censored observations, and the time interval from the start of the first treatment to the end of surgery (initiation of wound closure) was used as an estimate of the duration of transfusion-trigger reversal. The cumulative distribution of the primary efficacy variable of each treatment group was estimated by using the method of Kaplan–Meier. 21 Because subjects were comparable at baseline (see results), log-rank tests were performed to compare the distribution of the primary efficacy variable between groups using a step-down multiple-test comparison approach. 22 The initial comparison was planned to be between the P1.8group and the AB group. If that comparison was statistically significant (one-sided test, based on a significance level of 0.025), the P0.9group was compared with the AB group. Comparisons between the perflubron emulsion groups and the COL group were performed exploratively.
Secondary efficacy variables included the proportion of subjects achieving complete reversal of their first transfusion triggers, the proportion of subjects requiring allogeneic blood transfusions and the units of allogeneic blood transfused. All secondary efficacy parameters were tested using the Fisher exact test (two-sided and based on a significance level of 0.05) and analysis of variance as appropriate.
Patient demographics and baseline characteristics, pre- and post ANH data, data before and after the first treatment, and allogeneic blood transfusion data were summarized for each treatment group. Continuous variables were compared among groups by analysis of variance and, if an overall significance was found, followed by the Scheffé multiple-comparison test. Categoric variables (e.g.  , percentages) were compared among and between groups by Fisher's exact tests. Data before and after the first treatment were compared within groups using paired t  tests.
Results 
Two-hundred ninety patients were formally screened. Of these, 117 could not be enrolled because they violated one or more of the inclusion and exclusion criteria. Among these were 10 patients with post-ANH PvO2values < 40 mmHg. In addition, there were 26 patients who never reached a first transfusion trigger. Thus, a total of 147 patients were randomized, 38 patients to the COL group, 38 to the P0.9group, 36 to the P1.8group, and 35 to the AB group. Seven patients (5%) could not be evaluated at the 28-day follow up (COL, 2; P0.9, 1; P1.8, 2; AB, 2) but were included in the analysis of transfusion-trigger reversal and oxygen dynamics.
Patient baseline characteristics, type of surgical procedure, duration of surgery and anesthesia, and the percentage and volume of blood from preoperative autologous donation were similar among treatment groups (table 1). Hemoglobin levels before preoperative hemodilution were similar in all groups (COL, 11.7 ± 1.4 g/dl; P0.9, 11.6 ± 1.3 g/dl; P1.8, 11.7 ± 1.7 g/dl; AB, 11.7 ± 1.6 g/dl). The duration of preoperative hemodilution to a target hemoglobin level of 9 g/dl was also similar in all four groups (P  = 0.09). During ANH, similar amounts of blood were removed with similar amounts of plasma expander replacement (table 2). This resulted in similar hemoglobin concentrations, hemodynamic parameters, and arterial and mixed venous blood-gas values after hemodilution (table 2). Times from skin incision to first transfusion trigger, hemoglobin concentration, and hemodynamics also were not different among treatment groups when reaching the first transfusion trigger (table 3).
Table 1. Patient Characteristics 
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Table 1. Patient Characteristics 
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Table 2. Post–Acute Normovolemic Hemodilution Characteristics 
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Table 2. Post–Acute Normovolemic Hemodilution Characteristics 
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Table 3. Data prior to and after First Treatment 
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Table 3. Data prior to and after First Treatment 
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With the first treatment, PCWP remained essentially unchanged (table 3). Hemoglobin concentration decreased in both perflubron emulsion and colloid groups because of ongoing surgical blood loss and colloid replacement and remained relatively unchanged in the AB group receiving a unit of blood collected during ANH. Mean arterial pressure was relatively stable in all groups but increased in the P1.8group. Pulmonary artery pressure was unchanged in the COL and P1.8groups and minimally increased in the P0.9and AB groups. Heart rate remained relatively stable in all groups. Cardiac index increased in the P0.9, P1.8, and AB groups but was stable in the COL group. Arterial oxygen partial pressure increased (by protocol) in the COL, P0.9, and P1.8groups but remained unchanged in the AB group. PvO2and mixed venous oxygen saturation increased in all groups, but these measurements in the AB group were significantly lower after the first treatment compared with all other groups. Calculated arterial oxygen content increased only in the P0.9and P1.8groups and remained unchanged in the COL and AB groups. Mixed venous oxygen content increased in all groups. Oxygen delivery index increased in the P0.9, P1.8, and AB groups but remained unchanged in the COL group. Oxygen consumption index remained unchanged in all groups, and oxygen extraction decreased in all groups (table 3).
Percentage of complete transfusion-trigger reversal, that is reversal of all transfusion triggers (when multiple transfusion triggers were present), was highest in the P1.8group, followed by the P0.9, COL, and AB groups (P  = 0.001). The differences between the P1.8and AB (P  < 0.001) and COL (P  = 0.014) groups were statistically significant. Reversal of low PvO2was more frequent in the COL, P0.9, and P1.8groups compared with the AB group (table 4). Reversal of hypotension, tachycardia, and high cardiac output was not significantly different among groups (table 4). Duration of transfusion-trigger reversal (primary endpoint) was longest in the P1.8group, followed by the P0.9, AB, and COL groups (fig. 2and table 4). The differences in duration of reversal between the P1.8and AB (P  = 0.014) and COL (P  < 0.001) groups and the difference between P0.9and COL (P  = 0.012) were statistically significant.
Table 4. Transfusion Trigger Reversal 
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Table 4. Transfusion Trigger Reversal 
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Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
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Tachycardia, high cardiac output, and a hemoglobin concentration < 6 g/dl occurred with similar incidences in all treatment groups as a second transfusion trigger (table 5). Hypotension occurred more often in the COL group (45%) than in the P1.8group (11%), and a PvO2< 38 mmHg was recorded more often in the AB group compared with all other treatment groups.
Table 5. Distribution of Second Transfusion Triggers 
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Table 5. Distribution of Second Transfusion Triggers 
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There were no perioperative deaths through postoperative day 28. Allogeneic blood transfusion requirements were comparable in all treatment groups, and hemoglobin values at hospital discharge were similar (table 6).
Table 6. Allogeneic Blood Transfusion Data 
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Table 6. Allogeneic Blood Transfusion Data 
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No patient discontinued the study prematurely because of an adverse event. A total of 12 (COL, 6; P0.9, 2; P1.8, 2; AB, 2) patients experienced serious adverse events, all of which were assessed by the investigators as unlikely to be related to study treatment. A summary of adverse events is presented in table 7. The proportions of patients with any adverse events were 61% in the COL group, 53% in the P0.9group, 69% in the P1.8group, and 66% in the AB group. The majority of these were mild or moderate in severity, with no significant differences among groups. Platelet, bleeding, and clotting disorders were reported more frequently as adverse events in the P0.9(11%; n = 4) and P1.8(28%; n = 10) groups than in the COL (5%; n = 2) and AB (6%; n = 2) groups. These 14 adverse events in the perflubron emulsion groups included five cases of mild to moderate thrombocytopenia on the day of surgery, one case of thrombocytopenia (80,000/μl) on postoperative day 2, three cases of surgically induced bleeding, two cases of less intraoperative bleeding than expected, one elevated platelet count on postoperative day 13, and two cases with low coagulation parameters.
Table 7. Adverse Events Reported by ≥5% of Subjects by Body System and Preferred Term 
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Table 7. Adverse Events Reported by ≥5% of Subjects by Body System and Preferred Term 
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Postoperatively, platelet counts decreased in all groups as compared with baseline (fig. 3). An additional temporary decrease in platelet count between postoperative days 2 and 4 was observed in the P1.8group compared with the AB, COL, and P0.9groups (fig. 3). Platelet counts also were lower in the P0.9group on postoperative days 2 and 3 compared with the COL group, whereas no difference from the AB group was detected. No differences among groups were observed from postoperative day 7 onward.
Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
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Routine laboratory parameters were similar during the study period of 28 days in all treatment groups (data not shown). Mean duration of hospitalization was also similar across groups (15–17 days;table 1).
Discussion 
This study demonstrates that infusion of a 1.8-g/kg dose of perflubron emulsion combined with ventilation with 100% oxygen was well tolerated in all patients and resulted in hemodynamics and oxygen delivery changes similar to those found with blood transfusions. There was a higher frequency of “reversal of transfusion trigger” and a longer duration of reversal, although these conclusions are the results of relatively small differences in various parameters.
Avoiding allogeneic blood transfusions has become an important issue in the perioperative care of surgical patients 19,23 because of a variety of risks and side effects such as transfusion reactions, 1 alloimmunization, 2 transmission of infectious agents, 3,4 and immunosuppression. 5 As a result, there is a growing number of patients demanding perioperative treatment without allogeneic blood transfusions. Thus, in recent years a variety of methods and procedures have been developed to minimize allogeneic blood transfusions. Artificial oxygen carriers such as modified hemoglobin solutions or perfluorocarbon emulsions 24 could supplement these methods by providing additional oxygen delivery and oxygen unloading capacity and thereby enhancing patient safety during periods of intraoperative anemia.
The efficiency of perflubron emulsion in providing additional oxygen delivery and, more importantly, enhancing oxygen unloading capacity has been shown in experimental models of hemodilution and surgical blood loss 16,25 and cardiopulmonary bypass. 26 In these studies, animals treated with perflubron emulsion had a higher level of oxygen delivery because of an increased arterial oxygen content and the ability to maintain an increased cardiac output at low endogenous hemoglobin levels of 3 to 5 g/dl. 16,27,28 The increased oxygen delivery enhanced tissue oxygenation 25,29–31 and mixed venous oxygenation 16,26,27 and tended to improve survival after cardiopulmonary bypass 26 and after hemorrhagic shock with hypotensive resuscitation. 28 
Reversal of low mixed venous oxygen partial pressure in the present study in patients treated with perflubron emulsion and ventilation with an FIO2of 1.0 (table 4) is in agreement with recent experimental data. 16,25,31,32 Habler et al.  25 found that mixed-venous oxygen partial pressure was higher in animals treated with perflubron emulsion after hemodilution to a hemoglobin of 3 g/dl than in control animals, and parameters of left-ventricular contractility were found to be improved at a hemoglobin level of 3 g/dl after perflubron emulsion administration. 31 This may be the result of augmented oxygen delivery through very narrow microvascular channels that are more readily perfused with the tiny perflubron emulsion particles (< 0.2 μm in diameter) than relatively large red blood cells (7–8 μm in diameter). 33 Enhanced convective oxygen transport may increase local tissue oxygenation, particularly in the myocardium, 34 and thus contribute to maintaining blood pressure despite profound hemodilutional anemia. 25,27,31 
An increase in PvO2after the administration of perflubron emulsion was demonstrated previously in a pilot study in seven surgical patients. 35 The present study demonstrates that the increased oxygen delivery to the peripheral tissues seems to have a clinically relevant physiologic consequence, namely, the reversal of transfusion triggers (fig. 2and table 4), although a similar calculated amount of additional oxygen-carrying capacity was provided by each treatment (COL, 0.78 ml/dl; P0.9, 0.97 ml/dl; P1.8, 1.16 ml/dl; AB, 1.21 ml/dl; see Appendix 2). This provides evidence of efficient oxygen unloading by the perflubron emulsion. Interestingly, the duration of trigger reversal was the longest with administration of high-dose perflubron emulsion (fig. 2). This is particularly noteworthy because with ongoing surgical blood loss and concomitant volume replacement to maintain normovolemia a significant portion of the administered perflubron emulsion was lost, whereas in the colloid and 100% oxygen ventilation groups the full potential of the treatment was preserved. This suggests that the treatment benefit originated from the administration of perflubron and not primarily from the increased FIO2or the volume effect of the administered colloid.
We are aware of the complexity of using physiologic parameters such as individual tachycardia, hypotension, and low PvO2to indicate the need for a blood transfusion in anesthetized patients undergoing major surgery with significant blood loss. For this reason, a strict anesthesia and volume-replacement scheme was followed to differentiate, as much as possible, the manifestations of too much or too little anesthesia or hypovolemia from signs of compromised oxygen delivery resulting from normovolemic anemia. Thus, normovolemia was strictly mandated during the entire surgical procedure. In each patient normovolemia was achieved before and after ANH by an experienced anesthesiologist. The PCWP measured after ANH served as the best evidence of normovolemia and, by protocol, PCWP was maintained at or above this level during the entire surgical procedure. Evidence that this was indeed achieved included PCWP values 1–3 mmHg higher before the first treatment (table 3) than after ANH (table 2). Because normovolemia seems to have been well maintained during the operation, hypovolemia can be excluded as an important factor influencing the physiologic transfusion triggers.
The question remains whether the depth of anesthesia could be responsible for reaching physiologic transfusion triggers. We do not believe that this was likely, based on the anesthesia technique used and the fact that investigators were instructed to first assess and adjust depth of anesthesia if a physiologic transfusion trigger occurred. Anesthesia consisted of isoflurane, fentanyl, and vecuronium with ventilation with oxygen-enriched air without nitrous oxide. No epidural or spinal anesthesia was allowed, either as a primary anesthesia or as an adjunct to general anesthesia. Investigators were instructed to initially treat tachycardia with additional fentanyl, and hypotension with a reduction of isoflurane. Only if anesthesia adjustments to correct these changes were unsuccessful were they considered to be physiologic transfusion triggers.
The transfusion triggers used in this study can be criticized as being too liberal, that is, not representing critical values. However, for the purpose of this study, the exact values selected for the individual transfusion triggers are not critical to the evaluation of the drug's activity or efficacy of the four treatments because transfusion triggers were the same in all groups, and patients were randomized only after the first transfusion trigger was reached. More importantly, all groups were essentially identical at the point at which they reached the first transfusion trigger (table 3). However, the effects of the four treatments were different (fig. 2and tables 3 and 4).
Particularly important is the finding that duration of transfusion-trigger reversal in the high-dose perflubron emulsion group was the longest despite this group having the highest incidence of censored observations (table 4). Censored observations occurred in patients who did not reach a second transfusion trigger before the end of surgery. In 53% of patients treated with 1.8 g/kg perflubron emulsion at the first transfusion trigger, the time difference between the start of first treatment and end of surgery had to be used as an estimate for the duration of trigger reversal rather than the true duration, which always would have been longer. In addition, a hemoglobin level < 6 g/dl was set by the protocol as a second transfusion trigger. Hence, patients with no physiologic transfusion triggers who reached a hemoglobin level < 6 g/dl had to be transfused with one unit of autologous blood, thereby truncating the potential duration of transfusion-trigger reversal further. Both factors result in an underestimation of the total potential duration of transfusion-trigger reversal in the high-dose perflubron emulsion treatment group, because censored observations occurred significantly more frequently in this group compared with the COL and AB control groups (table 4).
Hypotension was recorded as a second transfusion trigger (table 5) more often in the COL group than in the P1.8group, the only treatment group in which an increase in mean arterial pressure was observed resulting from the first treatment (table 3). A low mixed venous oxygen tension occurred most often in the AB group, the group in which the lowest PvO2was measured after the first treatment. Distribution of second transfusion triggers therefore mirrored the initially observed treatment effect on physiologic parameters.
Allogeneic blood transfusion requirements were comparable among treatment groups (table 6). It must be stressed that this was not the primary goal of this study, and the design of the study essentially precluded the showing of any significant differences in transfusion outcome, because patients from all groups were managed at similar hemoglobin levels throughout the perioperative period. Hemoglobin levels were nearly identical at screening (table 1), before and after ANH (table 2), when reaching the first transfusion trigger (table 3), and after the first treatment (table 3), and patients in all groups were transfused to reach a hemoglobin level ≥ 8 g/dl after initiation of wound closure and were ultimately discharged from the hospital with similar hemoglobin levels (table 6). If there is bleeding at similar hemoglobin levels, it is to be expected that similar amounts of red blood cells will be lost, resulting in similar requirements for allogeneic blood transfusions. To show a reduction in allogeneic transfusion requirement requires future pivotal studies in which patients treated with perflubron emulsion are taken to lower hemoglobin levels to minimize the loss of red blood cells during surgical blood loss.
In conscious subjects, a delayed febrile response (6–8 h after dosing) along with flu-like symptoms and a moderate decrease in platelet count (3–4 days after dosing) have been described as side effects of perflubron administration. 33,36 In the present study, febrile responses were observed in 5 of 74 patients, compared with 1 of 73 in the groups not treated with perflubron emulsion (table 7). All other categories of adverse events were reported with similar incidences among treatment groups. Postoperative platelet counts were lower in perflubron emulsion–treated patients in the present study, with full recovery by postoperative day 7 (fig 3). This was not associated with any detectable adverse clinical event. This effect on platelet counts should continue to be investigated in future studies, particularly if redosing is allowed or if higher doses of perflubron emulsion are administered.
In conclusion, this study indicates that administration of perflubron emulsion combined with 100% oxygen ventilation is at least as effective as autologous blood in reversing physiologic transfusion triggers and thereby may represent a safe temporary alternative to conventional blood transfusion.
References 
References 
Sazama K: Reports of 355 transfusion-associated deaths: 1976 through 1985. Transfusion 1990; 30:583–90
Walker RH: Special report: Transfusion risks. Am J Clin Path 1987; 88:374–8
Ward JW, Holmberg SD, Allen JR, Cohn DL, Critchley SE, Kleinman SH, Lenes BA, Ravenholt O, Davis JR, Quinn MG, Jaffe HW: Transmission of human immunodeficiency virus (HIV) by blood transfusions screened as negative for HIV antibody. N Engl J Med 1988; 318:473–8
Schreiber GB, Busch MP, Kleinman SH, Korelitz JJ: The risk of transfusion-transmitted viral infections: The Retrovirus Epidemiology Donor Study. N Engl J Med 1996; 334:1685–90
Landers DF, Hill GE, Wong KC, Fox IJ: Blood transfusion–induced immunomodulation. Anesth Analg 1996; 82:187–204
Heiss MM, Mempel W, Jauch KW, Delanoff C, Mayer G, Mempel M, Eissner HJ, Schildberg FW: Beneficial effect of autologous blood transfusion on infectious complications after colorectal cancer surgery. Lancet 1993; 342:1328–33
Murphy P, Heal JM, Blumberg N: Infection or suspected infection after hip replacement surgery with autologous or homologous blood transfusions. Transfusion 1991; 31:212–7
Healy JC, Frankforter SA, Graves BK, Reddy RL, Beck JR: Preoperative autologous blood donation in total-hip arthroplasty: A cost-effectiveness analysis. Arch Pathol Lab Med 1994; 118:465–70
Blumberg N, Heal J, Chuang C, Murphy P, Agarwal M: Further evidence supporting a cause and effect relationship between blood transfusion and earlier cancer recurrence. Ann Surg 1988; 207:410–5
A report by the American Society of Anesthesiologists Task Force on Blood Component Therapy: Practice guidelines for blood component therapy. ANESTHESIOLOGY 1996; 84:732–47
American College of Physicians: Practice strategies for elective red blood cell transfusion. Ann Intern Med 1992; 116:403–6
Consensus conference: Perioperative red blood cell transfusion. JAMA 1988; 260:2700–3
Etchason J, Petz L, Keeler E, Calhoun L, Kleinman S, Snider C, Fink A, Brook R: The cost effectiveness of preoperative autologous blood donations. N Engl J Med 1995; 332:719–24
Bryson GL, Laupacis A, Wells GA: Does acute normovolemic hemodilution reduce perioperative allogeneic transfusion? A meta-analysis. The International Study of Perioperative Transfusion. Anesth Analg 1998; 86:9–15
Weiskopf RB: Mathematical analysis of isovolemic hemodilution indicates that it can decrease the need for allogeneic blood transfusion. Transfusion 1995; 35:37–41
Keipert PE, Faithfull NS, Bradley JD, Hazard DY, Hogan J, Levisetti MS, Peters RM: Oxygen delivery augmentation by low-dose perfluorochemical emulsion during profound normovolemic hemodilution. Adv Exp Med Biol 1994; 345:197–204
Vlahakes GJ, Lee R, Jacobs EE, LaRaia PJ, Austen WG: Hemodynamic effects and oxygen transport properties of a new blood substitute in a model of massive blood replacement. J Thorac Cardiovasc Surg 1990; 100:379–88
Zuck TF, Riess JG: Current status of injectable oxygen carriers. Crit Rev Clin Lab Sci 1994; 31:295–324
Spahn DR, Leone BJ, Reves JG, Pasch T: Cardiovascular and coronary physiology of acute isovolemic hemodilution: A review of nonoxygen-carrying and oxygen-carrying solutions. Anesth Analg 1994; 78:1000–21
Riess JG, Keipert PE. Update on perfluorocarbon-based oxygen delivery systems, in Blood Substitutes: Present and Future Perspectives. Edited by E. Tsuchida. Lausanne, Elsevier Science SA, 1998, pp 9:1–102
Kaplan EL, Meier P: Nonparametric estimation from incomplete observation. J Am Stat Assoc 1958; 53:457–481
Tamahane AC, Dunnett CW, A. H: A comparison of multiple test procedures for dose findings. Biometrics 1996; 52:21–37
Rutherford CJ, Kaplan HS: Autologous blood donations: Can we bank on it? N Engl J Med 1995; 332:740–2
Scott MG, Kucik DF, Goodnough LT, Monk TG: Blood substitutes: Evolution and future applications. Clin Chem 1997; 43:1724–1731
Habler OP, Kleen MS, Hutter JW, Podtschaske AH, Tiede M, Kemming GI, Welte MV, Corso CO, Batra S, Keipert PE, Faithfull NS, Messmer KF: Hemodilution and intravenous perflubron emulsion as an alternative to blood transfusion: Effects on tissue oxygenation during profound hemodilution in anesthetized dogs. Transfusion 1998; 38:145–55
Holman WL, Spruell RD, Ferguson ER, Clymer JJ, Vicente WV, Murrah CP, Pacifico AD: Tissue oxygenation with graded dissolved oxygen delivery during cardiopulmonary bypass. J Thorac Cardiovasc Surg 1995; 110:774–85
Cernaianu AC, Spence RK, Vassilidze TV, Gallueci JG, Gaprindashvili T, Olah A, Weiss RL, Cilley JH, Keipert PE, Faithfull NS, DelRossi AJ: Improvement in circulatory and oxygenation status by perflubron emulsion (Oxygent HT) in a canine model of surgical hemodilution. Artif Cells Blood Substit Immobil Biotechnol 1994; 22:965–77
Stern SA, Dronen SC, McGoron AJ, Wang X, Chaffins K, Millard R, Keipert PE, Faithfull NS: Effect of supplemental perfluorocarbon administration on hypotensive resuscitation of severe uncontrolled hemorrhage. Am J Emerg Med 1995; 13:269–75
Braun RD, Linsenmeier RA, Goldstick TK: New perfluorocarbon emulsion improves tissue oxygenation in cat retina. J-Appl-Physiol 1992; 72:1960–8
Mosca RS, Rohs TJ, Waterford RR, Childs KF, Brunsting LA, Bolling SF: Perfluorocarbon supplementation and postischemic cardiac function. Surgery 1996; 120:197–204
Habler OP, Kleen MS, Hutter JW, Podtschaske AH, Tiede M, Kemming GI, Welte MV, Corso CO, Batra S, Keipert PE, Faithfull NS, Messmer KF: Iv perflubron emulsion versus autologous transfusion in severe normovolemic anemia: Effects on left ventricular perfusion and function. Res Exp Med 1998; 197:301–18
Habler OP, Kleen MS, Hutter JW, Podtschaske AH, Tiede M, Kemming GI, Welte MV, Corso CO, Batra S, Keipert PE, Faithfull NS, Messmer KF: Effects of hyperoxic ventilation on hemodilution-induced changes in anesthetized dogs. Transfusion 1998; 38:135–44
Keipert PE: Perfluorochemical emulsions: Future alternatives to transfusion. Blood Subst Princ Meth Prod Clin Trials 1998; 2:127–156
Faithfull NS: Oxygen delivery from fluorocarbon emulsions: Aspects of convective and diffusive transport. Biomater Artif Cells Immobilization Biotechnol 1992; 20:797–804
Wahr JA, Trouwborst A, Spence RK, Henny CP, Cernaianu AC, Graziano GP, Tremper KK, Flaim KE, Keipert PE, Faithfull NS, Clymer JJ: A pilot study of the effects of a perflubron emulsion, AF 0104, on mixed venous oxygen tension in anesthetized surgical patients. Anesth Analg 1996; 82:103–7
Keipert PE, Faithfull NS, Roth DJ, Bradley JD, Batra S, Jochelson P, Flaim KE: Supporting tissue oxygenation during acute surgical bleeding using a perfluorochemical-based oxygen carrier. Adv Exp Med Biol 1996; 388:603–9
Appendix 1 
The European Perflubron Emulsion Study Group 
D. R. Spahn, Ch. Rippmann (Institute of Anesthesiology, University Hospital, Zurich, Switzerland); E. Vandermeersch, R. van Brempt, L. Veeckman, R. Vranckx (Department of Anesthesiology, University Hospital Leuven, Belgium); H. van Aken, G. Theilmeier, T. Prien, K. Singbartl, N. Mertes, C. Schneider (Department of Anesthesiology and Intensive Care Medicine, University Hospital Münster, Germany); G. Hempelmann, J. P. Reibold, P. von Rosen, K. Wulf (Department of Anesthesiology, University Hospital, Giessen, Germany); K. Messmer, M. Welte, O. Habler, A. Hagemann, M. Kleen (Institute for Surgical Research and Department of Anesthesiology, Grosshadern Hospital, Munich, Germany); S. de Lange, L. Berry, P. de Laat, F. A. Kwinten, C. D. Punt (Department of Anesthesiology, Academisch Ziekenhuis Maastricht, The Netherlands); P. Musto (Department of Anesthesiology, Leicester General Hospital, Leicester, U.K.); D. M. Albrecht, M. Ragaller, M. Gama de Abreu (Department of Anesthesiology and Intensive Care, University Hospital Dresden, Germany); K. van Ackern, K. Waschke, W. Segiet, T. Frietsch (Department of Anesthesiology and Intensive Care, Hospital Mannheim, Germany); E. W. G. Weber, R. Slappendeel, J. J. de Hoog (Department of Anesthesiology, Sint Martinskliniek, Nijmwegen, The Netherlands); F. Mercuriali, G. Inghilleri, A. d'Aloia, R. Coluccia (Istituto Ortopedico Gaetano Pini, Milano, Italy).
Appendix 2 
Equations to Compute Oxygen Dynamics 
Oxygen Content 
MATH 1where CaO2denotes arterial oxygen content (ml/dl), CaO2TOTALdenotes total CaO2, CaO2Hbdenotes hemoglobin-bound CaO2, CaO2PL is the plasma-contained CaO2, and CaO2PERFLis the perflubron emulsion transported oxygen level. MATH 2MATH 3where SaO2and SvO2denote arterial and mixed venous oxygen saturations [%]; PaO2and PvO2denote arterial and mixed venous oxygen partial pressures (mmHg);[PERFL] denotes perflubron concentration (g/dl); 1.34 is the oxygen carrying capacity of hemoglobin (ml O2/g Hb); 0.003 is the solubility coefficient of oxygen in plasma (ml O2/mmHg PO2); 0.53 is the solubility of oxygen (ml O2) in 1 ml perflubron at one atmosphere (760 mmHg) and 37°C; and 1.92 is the density of perflubron (g/ml).
Oxygen Consumption 
MATH 4where VO2Idenotes oxygen consumption index and CI denotes cardiac index (cardiac output/body surface area)(l · min−1· m−2).
Oxygen Delivery 
MATH 5where DO2Idenotes oxygen delivery index.
Oxygen Extraction 
MATH 6where O2Exoveralldenotes overall oxygen extraction ratio (%). MATH 7where O2Exhemoglobindenotes hemoglobin oxygen extractionratio (%).
These formulae do not take into account the small volume of the fluorocrit after perflubron emulsion application. However, fluorocrit was not measured, and the theoretically expected correction factor is 1.2 ± 0.4% for a 1.8-g/kg perflubron emulsion dose.
To compute the theoretical amount of additional oxygen carrying capacity initially offered by the four treatments the following assumptions were made: a pretreatment arterial oxygen partial pressure of approximately 160 mmHg at an FIO2of 0.4 and a posttreatment pressure of approximately 420 mmHg at an FIO2of 1.0 (except for the AB group)(table 3), a body weight of 70 kg, an estimated blood volume of 5,000 ml, and a hemoglobin concentration of 10 g/dl (with a protocol-defined ANH hemoglobin target of 9 g/dl).
Perflubron concentration in whole-blood samples was dertermined by gas-chromatographic headspace analysis, in which a sample of the gas phase from a closed blood sample vial is injected and analyzed by gas chromatography and then compared with a standard curve generated by spiking whole blood with known quantities of perflubron emulsion.
Fig. 1. Study protocol. 
Fig. 1. Study protocol. 
Fig. 1. Study protocol. 
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Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
Fig. 2. Duration of reversal due to treatment at first transfusion trigger. Treatments were infusion of 450 ml colloid and ventilation with an inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), and 450 ml autologous blood with FIO2of 0.4 (AB). 
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Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
Fig. 3. Postoperative platelet counts in patients treated with infusion of 450 ml colloid and ventilation with inspiratory oxygen fraction (FIO2) of 1.0 (COL), perflubron emulsion 0.9 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P0.9), perflubron emulsion 1.8 g/kg and colloid (total of 450 ml) with FIO2of 1.0 (P1.8), or 450 ml autologous blood with FIO2of 0.4 (AB). *  P  < 0.05  vs.  COL;†  P  < 0.05  vs.  AB. 
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Table 1. Patient Characteristics 
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Table 1. Patient Characteristics 
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Table 2. Post–Acute Normovolemic Hemodilution Characteristics 
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Table 2. Post–Acute Normovolemic Hemodilution Characteristics 
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Table 3. Data prior to and after First Treatment 
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Table 3. Data prior to and after First Treatment 
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Table 4. Transfusion Trigger Reversal 
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Table 4. Transfusion Trigger Reversal 
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Table 5. Distribution of Second Transfusion Triggers 
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Table 5. Distribution of Second Transfusion Triggers 
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Table 6. Allogeneic Blood Transfusion Data 
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Table 6. Allogeneic Blood Transfusion Data 
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Table 7. Adverse Events Reported by ≥5% of Subjects by Body System and Preferred Term 
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Table 7. Adverse Events Reported by ≥5% of Subjects by Body System and Preferred Term 
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