Transfusion Medicine in Veterinary Emergency and Critical Care Medicine Elizabeth Rozanski, DVM, DACVECC, DACVIM and Armelle M. de Laforcade, DVM, DACVECC
Transfusion medicine is a vital part of veterinary emergency and critical care medicine. The goals of this article are to review blood banking and the transfusion principles surrounding care of the critically ill or injured small animal, to highlight the differences in emergency/critical care transfusions compared with standard transfusion medicine, and to discuss traumatic blood loss and sepsis as unique entities in emergency and critical medicine. © 2004 Elsevier Inc. All rights reserved.
dequate transfusion resources are crucial to a successful patient outcome. As transfusion medicine has grown in recent years, so has the availability of commercial blood products, thus increasing the ability of many emergency hospitals to have immediate access to blood products and expert consultants. Emergency medicine relies upon preparation for potential life-threatening situations. This means that hospitals engaged in emergency medicine need to have a plan for transfusions prior to actual need. There are a variety of options available for meeting the potential transfusion needs of critically ill dogs and cats. A list of commercial blood banks and further information about blood banking and transfusion medicine may be found at the Association of Veterinary Hematology and Transfusion Medicine’s Web site (http://www.vetmed.wsu.edu/org-avhtm/ index.asp). However, it is essential that an active emergency program have sufficient blood, plasma, or blood substitutes on hand. Canine transfusions are commonly provided as components, such as packed red blood cells (pRBCs) and fresh-frozen plasma (FFP) in order to best use available resources.1-4 Each unit (⬃450 mL) of whole blood, if processed promptly, may be divided into 1 unit of pRBCs and 1 unit of FFP. FFP has a shelf-life of 1 year, after which it is stable for an additional 4 years as frozen plasma. Stored plasma is still useful for treating anticoagulant rodenticide toxicities and for supplemental albumin. FFP may also be divided into cryoprecipitate, which contains increased concentrations of factor VIII, von Willebrand’s factor, fibrinogen, and cryo-poor plasma. Cryo-poor plasma is inappropriate for administration to patients with von Willebrand’s disease and hemophilia. Fresh whole blood may also be processed into platelet-rich plasma or platelet concentrates, although this is rarely done in veterinary medicine. pRBCs
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From the Section of Critical Care, Department of Clinical Sciences, Tufts University, North Grafton, MA, USA. Address reprint requests to Dr. Elizabeth Rozanski, Section of Critical Care, Department of Clinical Sciences, Tufts University, 200 Westboro Rd., North Grafton, MA 01536. E-mail: Elizabeth.rozanski@tufts.edu © 2004 Elsevier Inc. All rights reserved. 1096-2867/04/1902-0005$30.00/0 doi:10.1053/j.ctsap.2004.01.005
typically have a shelf life of 30 to 35 days at most. Additive solutions (eg, Nutricel and Adsol) are available that can extend shelf life to 35 to 37 days. Each single unit of blood processed represents a technical time commitment of approximately 30 to 45 minutes, not including the time needed to recruit and screen eligible donors. Most canine donors are very tolerant of the donation process and, in fact, many commercial blood banks actively exclude dogs that do not appear to be willing donors. Blood products may be purchased from commercial blood banks or collected and processed in-house before emergent need. The once common practice of bleeding a staff member’s personal dog or an in-house donor on an as-needed basis is not appropriate for most emergency practices because of time constraints and the manpower required. Because pRBCs have a relatively short lifespan and emergency needs are notoriously unpredictable, it is a challenge to decide the amount of pRBCs to keep on hand. It would appear prudent that practices should carefully monitor their blood banks and strive to maintain 25% to 50% more units that they typically use in a given month. Obtaining blood for feline transfusions is inherently more challenging than for canine blood transfusions. Cats’ blood volume is less than dogs’ (60 mL/kg vs. 90 mL/kg) and donation typically requires sedation. Additionally, cats have a wider assortment of clearly blood-borne diseases than dogs, such as feline leukemia virus, feline immunodeficiency virus, and Mycoplasma haemofelis/haemominutum (formerly known as Hemobartonella felis). A recent abstract documented the prevalence of infection in the blood-donor pool.5 Cat blood transfusions are commonly administered as whole blood (either fresh or stored) in part because of the challenge of preparing components from a small volume of blood. Whole blood can be separated into pRBCs and FFP for patients requiring these specific components. Autotransfusion may also be a viable option in certain circumstances.6,7 Autotransfusion relies upon the collection and subsequent re-infusion of the patient’s own shed blood. Autotransfusion has been useful in situations where blood is not available in a timely fashion. Additionally, autotransfusion is particularly appealing because there is obviously no chance of an immunological reaction to one’s own red cells. Autotransfusion is performed by collecting blood lost into either the thoracic or abdominal cavity and then filtering and re-infusing the blood. Blood is collected into either sterile fluid-administration bags or blood-collection sets. The blood is commonly collected with an anticoagulant, although this may not be required if the effusion has been present for several hours. Standard transfusion-administration sets with filters should be used to minimize the transfusion of clots and small debris. Autotransfusion is contraindicated if contamination with urine, bacteria, or bile is possible, and relatively contraindicated in the presence of diffuse neoplasia. Because of the recent growth in blood banking
Clinical Techniques in Small Animal Practice, Vol 19, No 2 (May), 2004: pp 83-87
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and the improved availability of a wide variety of blood products, the need for autotransfusion has declined in recent years. However, it remains a useful alternative in emergency practice. The recent development and marketing of purified bovine hemoglobin (OxyglobinÂŽ, Biopure, Cambridge, MA), which may be used as a blood substitute or as a colloidal solution that carries oxygen, has provided a valuable product for emergency and critical care medicine. Oxyglobin has been approved for the treatment of anemia only in dogs, but its use has also been reported in cats.8 The recommended doses for dogs are 10 to 30 mL/kg. There is not a published dose for cats, although a dose of 5 to 15 mL/kg is used clinically. Cats are prone to volume overload, particularly when transfused with a colloid. Therefore, careful monitoring of respiratory rate/effort, and possibly central venous pressure, is warranted whenever oxyglobin is used in cats. Hemoglobin-based oxygen carriers also have some potential theoretical advantages in the therapy of shock because various rheological characteristics of purified hemoglobin may improve oxygen delivery to various sites.8-10 Additionally, oxyglobin has a long shelf life and no special storage requirements. Thus, the addition of oxyglobin to the blood bank of an emergency/critical care facility is beneficial, particularly in circumstances where blood needs are variable.
Indications for Transfusions in Emergency and Critical Care Indications for transfusions in emergency and critical care practice are often similar to those in surgery or internal medicine, and include most commonly anemia and coagulopathy. To best understand the type and rate of blood transfusion indicated, Anemia can be divided into normovolemic anemia and hypovolemic anemia.
Normovolemic Anemia Animals that are anemic with a normal intravascular volume status typically have either non-regenerative or hemolytic causes for their anemia. In these cases, the anemia may have been long-standing and/or accompanied by a relative increase in plasma volume. Normovolemic anemia is better tolerated by animals and requires a less urgent treatment. Normovolemic anemic patients are clinically appreciated by appearing relatively bright on physical examination, having a normal or only marginally elevated heart and respiratory rate, having normal to slightly elevated total solids (typically greater the 5.8g/dL), and having pale pink rather than white mucous membranes. Blood pressure should be normal, although there may be a large pulse pressure, reflecting increased stroke volume and possibly diastolic run-off. The decision to transfuse these patients will typically occur at a much lower packed cell volume (PCV) than for hypovolemic patients. Transfusion should be considered when clinical signs attributable to anemia are present (ie, tachypnea, tachycardia, weakness) or when an intervention (eg, surgery) is contemplated. pRBCs would be the component of choice for administration to normovolemic anemic patients because of its lower oncotic pressure. Also, diluent volume can be decreased to enable delivery of the same number of RBCs with a smaller total volume. Blood products should be administered at a slower rate in these patients to minimize the risk of volume overload. This may necessitate division of units into smaller aliquots and refriger-
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ation until administration to minimize the risk of bacterial contamination. The target hematocrit after transfusion in a normovolemic anemia patient accompanying either hemolysis or non-regeneration is typically in the low 20s. Formulas (eg, 1 mL/kg of pRBC will raise PCV âŹƒ1%) to calculate the required transfusion volume are usually accurate in these patients.
Hypovolemic Anemia In comparison, hypovolemic anemic patients have typically suffered from catastrophic blood loss within the last few hours.11,12 These patients are very weak, and have rapid and faint pulses, with their mucous membranes appearing whiter than their hematocrit might suggest. Their total solids are low, corresponding with the loss of red-cell mass, although early on there may be no change seen. These patients require aggressive volume resuscitation as well as blood transfusion. Some patients may require surgical intervention to control hemorrhage. It is not uncommon to find that the post-transfusion hematocrit may be lower than the starting point because of dilution of the circulation volume with crystalloids or colloids, as well as ongoing hemorrhage. However, clinical improvement is seen (ie, normalization of heart rate, blood pressure) despite the drop in hematocrit. It is crucial for the clinician to remember that oxygen delivery may be improved dramatically by increasing blood volume (and thus improving cardiac output), even in the face of a lower absolute number for the hematocrit. No formula exists that may predict the volume of blood required to resuscitate these patients adequately. In nonemergency situations, transfusions are typically administered over 4 hours. It may be necessary to infuse blood products more rapidly, with a pressure bag or rapid infuser. It is important to establish that the administration pumps being used are safe for blood administration because damage to the RBCs can cause hemolysis. A recent retrospective study described the clinical outcome in dogs experiencing massive transfusion, and documented predictable changes in electrolytes and coagulation status.12 Massive transfusion is defined as a transfusion with blood or blood components (pRBCs or FFP) of an entire blood volume (90 mL/kg) in 24 hours or less, or transfusion of half a blood volume in 3 hours. Animals receiving massive transfusions may develop electrolytes disturbances, thrombocytopenia, and dilutional coagulopathy. The most common electrolyte abnormality seen with massive transfusion is hypocalcemia. The anticoagulant citrate acts by binding calcium and thus prevents the progression of the coagulation cascade. When large amounts of citrate are administered, the liver’s ability to metabolize it is overwhelmed. This leads to chelation of calcium in the recipient. Clinically significant hypocalcemia may result in tremors or hypotension. Infusion of calcium gluconate may be warranted (30 mg/kg intravenously over 2-5 minutes) in critically ill patients with massive transfusion and hypotension. Transfusion of a large volume of pRBCs may also result in the dilution of coaguation factors and platelets, resulting in thrombocytopenia and dilutional coagulopathy. Plasma transfusions are indicated to treat coagulopathy associated with decreased or malfunctioning coagulation factors.1,13-15 Coagulopathies may be inherited or acquired in emergency patients. Coagulopathies are typically diagnosed based on prolongation of standard coagulation tests [prothrombin time (PT), activated partial thromboplastin time (aPTT), activated clotting time (ACT)]. Because of the reserve of the ROZANSKI AND DE LAFORCADE
coagulation system, a large percentage of the clotting factors needs to be lost before prolongation of the standard coagulation tests. The results of these assays are also affected by plasma pH and body temperature. However, because the laboratory tests are performed at standard temperatures (37°C), the effect of hypothermia may be undetectable in vitro. Normalization of these variables during testing may influence results ex vivo. It is not uncommon for critically ill patients to have normal coagulation parameters yet continue to hemorrhage. FFP transfusion will not clearly normalize vague coagulopathy. Point-of-care analyzers have been evaluated for the assessment of PT, aPTT, and ACT in dogs.16 This enables the immediate assessment of coagulation, whereas a sample sent out to a laboratory may no longer reflect the patient’s current status because of the delay in receiving the results. Significant prolongations (⬎25% control value) of PT and aPTT should be treated aggressively with supportive care for the underlying disease, plasma, and/or supplemental vitamin K. Typically, in human medicine, plasma transfusion is not considered unless hemorrhage is present in the face of prolonged coagulation testing. Vitamin K is particularly warranted in coagulopathy associated with anticoagulant rodenticides and liver failure because activation of vitamin K-dependent coagulation factors (II, VII, IX, and X) is impaired in these patients. Acquired coagulopathies are common in critically ill animals and include underlying causes such as anticoagulant rodenticide toxicities, sepsis with and without disseminated intravascular coagulation (DIC), neoplasia, liver failure, and over-heparinization.17 Inherited coagulopathies are far less common in emergency and critical care settings. Inherited coagulopathies are typically identified after abnormal hemorrhage in puppy or kittenhood. However, occasionally the emergency clinician is presented with a previously undiagnosed case that is currently bleeding. Inherited coagulopathies that may benefit from plasma transfusion include hemophilia A (factor VIII deficiency) or B (factor IX deficiency), von Willebrand’s disease (von Willebrand’s factor deficiency), and specific isolated coagulation-factor deficiencies. Patients with hemophilia A or B will often present with excessive hemorrhage after minor trauma. These patients will have a prolonged aPTT but a normal PT. Confirmation of a diagnosis of these disorders relies on performing specific factor VIII or factor IX assays. Because they cannot be differentiated clinically, both assays should be performed when suspected.18 Patients with von Willebrand’s disease will typically present with mucosal hemorrhage, skin bruising, and prolonged bleeding from wounds. Coagulation tests in these patients are normal because von Willebrand’s factor is actually involved in primary hemostasis. Prolongation of buccal mucosal bleeding time in the face of a normal platelet count increases the index of suspicion for this disease. Confirmation of a diagnosis requires von Willebrand’s factor measurement. If a patient presents with a suspected inherited coagulopathy, then it is important to collect and freeze citrated plasma samples for possible factor levels before transfusion. Transfusions may be indicated as a therapy in patients with sepsis/multiple organ dysfunction syndrome, pancreatitis, hypoalbuminemia, and DIC without associated laboratoryproven coagulopathy. It appears commonplace among emergency and critical care clinicians to want to give plasma when animals appear critically ill. However, it is prudent to try to determine what is the desired endpoint of transfusion (ie, norTRANSFUSION IN VETERINARY EMERGENCY AND ICU MEDICINE
malization of a specific laboratory parameter or cardiovascular parameter). Correction of an albumin deficit is usually impractical in all but very small patients because much of the patient’s albumin is actually located extravascularly. Recently, the use of human albumin has been described in critically ill animals; however, no definitive guidelines have been published. Additionally, as a human product, there exists the potential for human albumin to produce antigen stimulation or other adverse reactions in veterinary patients.
Trauma Specific guidelines for transfusion medicine in veterinary critical care require individual assessment of each patient. Trauma with subsequent hemorrhage is a common indication for transfusion.11 In dogs, traumatic hemorrhage frequently results from intra-abdominal (liver/spleen) damage, intra-thoracic trauma, or bleeding into a fracture site.3,11 Management of severely affected dogs may be particularly challenging. Different authors may advise different approaches. Specifically, for intra-abdominal hemorrhage, management may be surgical or medical. Surgical management involves fluid resuscitation followed by rapid celiotomy with control of hemorrhage.11 Advantages of surgical management include the ability to evaluate individual organs for damage, including inspection of the mesentery and intestinal vasculature. Additionally, rapid control of hemorrhage may be helpful in preventing the complications associated with massive transfusion, such as dilutional coagulopathy, thrombocytopenia, and hypoproteinemia. Disadvantages of surgery include the additional stresses on the system that general anesthesia and a celiotomy may bring. Additionally, the financial impact of surgery can be large, although it may actually be less of a financial burden to the clients if the length of hospitalization and/or number of transfusions required can be reduced. Medical management of traumatic hemoabdomen includes the application of a tight abdominal wrap, including the entire abdomen, and intravascular support with blood transfusions and crystalloids and/or colloids. An abdominal wrap should not be applied if the patient shows signs of respiratory compromise. Preference for medical or surgical management of traumatic hemoabdomen is often variable between institutions and individual emergency clinicians. In one retrospective study, no survival difference was found between operated and non-operated dogs with traumatic hemoabdomen.11 In the human medical literature, a discussion of traumatic shock is often accompanied by the term “delayed” or “hypotensive” resuscitation.19-22 Delayed or hypotensive resuscitation refers to withholding large volumes of resuscitative fluids until surgical control of hemorrhage is possible. This concept originated in the field of human trauma medicine when early surgical intervention was shown to accompany better survival. Thus, in a critically injured person, fluid resuscitation would be titrated to maintain a minimally acceptable systolic blood pressure (⬃60 mm Hg) until the source of the hemorrhage (eg, splenic laceration, pelvic fracture) could be definitively treated. A large body of evidence shows that in both the research and the clinical setting systolic blood pressure does not need to be returned completely to normal to maximize patient survival.19,22 The rationale for this approach is that restoration of normal blood pressure may actually result in increased hemorrhage through increased blood pressure. One study docu-
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mented an improvement in survival for human patients with penetrating thoracic injuries who underwent delayed fluid resuscitation rather than immediate therapy.22 No studies have evaluated delayed or hypotensive fluid resuscitation in veterinary patients, although a large number of studies have been performed in the research setting. It appears likely that hypotensive or delayed fluid resuscitation may be in order in some critically injured dogs, although no guidelines yet exist. However, it is essential to remember that delayed resuscitation does not translate to no resuscitation, and critically injured animals should be stabilized appropriately.
Sepsis Transfusion therapy is also a crucial part of therapy for animals with sepsis, neoplasia, and/or DIC.17 Hematological abnormalities, including leukopenia or leukocytosis, thrombocytopenia, anemia, and coagulopathy as defined by prolongation of PT and aPPT are very common in patients with sepsis.23,24 In patients with sepsis and anemia, red cells should be transfused in order to maintain a hematocrit sufficient for oxygen delivery. Oxygen delivery is determined by the following formula: Oxygen content ⫻ cardiac output. Oxygen content is equal to (hemoglobin ⫻ 1.34 ⫻ SPO2) ⫹ (PaO2 ⫻ 0.003), and cardiac output is equal to heart rate ⫻ stroke volume. The target hematocrit recommended to assure adequate tissue oxygenation has not been defined in animals. In people, recommendations are to maintain the hemoglobin at values equal to or greater than 7 gm/dL (a Hemocrit of ⬃21%). One retrospective study of 74 dogs receiving FFP transfusion documented that sepsis was an indication for transfusion in many dogs.2 Potential benefits of plasma in sepsis include replacement of depleted procoagulant factors, antithrombin (AT), and albumin. AT is important in sepsis as a naturally occurring anticoagulant protein responsible for 70% to 80% of the anticoagulant activity of the plasma. In sepsis (and DIC), AT is consumed at a high rate, and may predispose a patient to thrombotic disease. One small prospective study did not document an increase in AT 24 hours after transfusion in critically dogs.25 In critically ill people, plasma-transfusion recommendations for sepsis are variable and support normalization of the coagulogram.13,14 The dose of plasma indicated for patients with sepsis is poorly defined. Interestingly, a recent study in people documented the relative failure of FFP infusions to normalize PT in chronic liver disease at clinically common doses, and recommendations include increasing the units transfused from 2-4 to 6 or greater.15 As with liver disease, patients with DIC may have a continuous depletion of coagulation factors that may require higher doses of plasma than a patient for which the factor loss is finite (eg, a dog with rodenticide intoxication that is subsequently treated with vitamin K). In any case, it is certainly prudent not to transfuse plasma to replace depleted factors without careful monitoring of the effect on PT and aPPT. Activated protein C is the only preparation that has been documented to improve survival in people with sepsis.26,27 This is a recombinant product designed for human use, and the role and associated cost of treatment in animals with activated protein C is unknown at this point. It is extrapolated that at the time of this writing (fall 2003), the cost to treat an average-size large-breed dog with activated protein C would be in excess of $5,000.
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Conclusions Transfusion medicine is, and will continue to be, an important area of emergency and critical care medicine. Transfusions, including pRBCs, FFP, and hemoglobin-based oxygen-carrying solutions, may be beneficial for a variety of reasons. However, it is wise to carefully consider the predicted benefit in each patient and then post-transfusion to assess whether that benefit was reached. Additional prospective clinical trials in specific areas associated with transfusions in emergency and critical care are warranted, because many current guidelines are based on opinion and conjecture rather than on evidence-based medicine.
References 1. Logan JC, Callan MB, Drew K, et al: Clinical indications for use of fresh frozen plasma in dogs: 74 dogs (October through December 1999). J Am Vet Med Assoc 218:1449-1455, 2001 2. Callan MB, Oakley DA, Shofer FS, et al: Canine red blood cell transfusion practice. J Am Anim Hosp Assoc 32:303-311, 1996 3. Kerl ME, Hohenhaus AE: Packed red blood cell transfusions in dogs: 131 cases (1989). J Am Vet Med Assoc 202:1495-1499, 1993 4. Lanevschi A, Wardrop KJ: Principles of transfusion medicine in small animals. Can Vet J 42:447-454, 2001 5. Hackett TB, Lappin MR, Jensen WA: Prevalence of Mycoplasma haemofelis and Mycoplasma haemominutum in blood donor cats. Proceedings of the American College of Veterinary Internal Medicine, Charlotte, North Carolina, 2003 6. Crowe DT: Autotransfusion in the trauma patient. Vet Clin North Am Small Anim Pract 10:581-597, 1980 7. Purvis D: Autotransfusion in the emergency patient. Vet Clin North Am Small Anim Pract 25:1291-1304, 1995 8. Gibson GR, Callan MB, Hoffman V, et al: Use of hemoglobin-based oxygen-carrying solution in cats: 72 cases (1998-2000). J Am Vet Med Assoc 221:96-102, 2002 9. Wohl JS, Cotter SM: Blood substitutes: oxygen-carrying acellular fluids. Vet Clin North Am Small Anim Pract 25:1417-1440, 1995 10. Muir WW, Wellman ML: Hemoglobin solutions and tissue oxygenation. J Vet Intern Med 17:127-135, 2003 11. Mongil CM, Drobatz KJ, Hendricks JC: Traumatic hemoperitoneum in 28 cases: a retrospective review. J Am Anim Hosp Assoc 31:217222, 1995 12. Jutkowitz LA, Rozanski EA, Moreau JA, et al: Massive transfusion in dogs: 15 cases (1997-2001). J Am Vet Med Assoc 220:1664-1669, 2002 13. Hellstern P, Mutean W, Schramm W, et al: Practical guidelines for the clinical use of plasma. Thromb Res 107:S53-S57, 2002 (suppl) 14. Mueller MM, Bomke B, Siefried E: Fresh frozen plasma in patients with disseminated intravascular coagulation or in patients with liver diseases. Thromb Res 107:S9-S17, 2002 (suppl) 15. Youssef WI, Salazar F, Dasarathy S, et al: Role of fresh frozen plasma infusion in correction of coagulopathy of chronic liver disease: a dual phase study. Am J Gastoenterol 98:1391-1394, 2003 16. Tseng LW, Hughes D, Giger U: Evaluation of a point-of-care coagulation analyzer for measurement of prothrombin time, activated partial thromboplastin time, and activated clotting time in dogs. Am J Vet Res 62:1455-1460, 2001 17. Hohenhaus AE: Transfusion issues in the cancer patient. Clin Tech Small Anim Pract 18:135-138, 2003 18. Mansell P: Hemophilia A and B, in Feldman BF, Zinkl JG, Jain NC (eds): Schalm’s Veterinary Hematology, ed 5. Philadelphia, PA, Lippincott Williams & Wilkins, 2000, pp 1026-1029 19. Varela JE, Cohn SM, Diaz I, et al: Splanchic perfusion during delayed, hypotensive, or aggressive fluid resuscitation from uncontrolled hemorrhage. Shock 20:476-480, 2003 20. Wade CE, Grady JJ, Kramer GC: Efficacy of hypertonic saline dextran fluid resuscitation for patients with hypotension from penetrating trauma. J Trauma 54:S144-S148, 2003 (suppl) 21. Stern SA: Low-volume fluid resuscitation for presumed hemorrhage shock: helpful or harmful? Curr Opin Crit Care 7:422-430, 2001 ROZANSKI AND DE LAFORCADE
22. Bickell WH, Wall MJ, Pepe PE, et al: Immediate versus delayed fluid resuscitation for hypotensive patients with penetrating torso injuries. N Engl J Med 331:1105-1109, 1994 23. Aird WC: The hematologic system as a marker of organ dysfunction in sepsis. Mayo Clin Proc 78:869-881, 2003 24. Strauss R, Wehler M, Mehler K, et al: Thrombocytopenia in patients in the medical intensive care unit bleeding prevalence, transfusion requirements and outcome. Crit Care Med 30:1765-1771, 2002 25. Rozanski EA, Hughes D, Giger U: The effect of heparin and fresh
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frozen plasma on antithombin III activity, prothrombin time and activated partial thromboplastin time in critically ill dogs. JVECCS 11:15-21, 2001 26. McCoy C, Matthews SJ: Drotrecogin alfa (recombinant human activated protein C) for the treatment of severe sepsis. Clin Ther 25:396421, 2003 27. Bernard GR, Vincent JL, Laterre PF, et al: Efficacy and safety of recombinant human activated protein C for severe sepsis. N Engl J Med 344:699-709, 2001
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