CHAPTER 142 PRESERVATION AND CLINICAL USE OF PLATELETS
CHAPTER 142 PRESERVATION AND CLINICAL USE OF PLATELETS
Techniques for Platelet Preparation
Whole-Blood-Derived Platelet Concentrates
Apheresis Platelet Concentrates
Storage of Platelet Concentrates
Liquid Storage at 20 to 24°C
Fresh Whole Blood
Lyophilized Platelets, Platelet Membranes, and Platelet Substitutes
General Principles in Patients with Marrow Failure
Platelet Transfusion Trigger
Thrombocytopenia Due to Platelet Loss, Sequestration, or Destruction
Immune (Idiopathic) Thrombocytopenic Purpura (ITP)
Neonatal Alloimmune Thrombocytopenia
Qualitative Platelet Disorders
Possible Contraindications to Platelet Transfusion
Complications of Platelet Transfusion
Complications Due to Contaminating Leukocytes
Alloimmunization to Class I HLA Antigens
Management of the Refractory Patient
Febrile Nonhemolytic Transfusion Reactions (FNHTR)
Transmission of Cytomegalovirus (CMV)
Graft-Versus-Host Disease (GVHD)
Complications of Filtration of Platelet Concentrates
Complications Due to Contaminating Red Cells
Complications Due to Plasma and its Contents
The use of platelet transfusions increased dramatically in the 1980s and peaked in 1992, following which, at least in the United States, it has remained stable. Worldwide, many methods are used for the preparation of platelets for transfusion. The “platelet-rich plasma” method and the “buffy coat method” are popular for the separation of platelets from whole blood donations in North America and in Europe, respectively. In addition, platelets separated by apheresis are gaining in popularity worldwide in order to limit donor exposure and to minimize the number of contaminating leukocytes in the preparations. Many institutions are making their platelet products universally leukoreduced at the time of their preparation. After preparation, platelets are generally stored at 20 to 24°C within plastic containers whose walls are adequately permeable to oxygen. It is optimal to agitate these preparations continuously. Storage at lower temperatures results in decreased in vivo survival after transfusion, and adequate access to oxygen and agitation are required to prevent deleterious declines in pH. Platelets stored in this fashion produce satisfactory clinical responses after storage for 5 to 7 days. Currently, storage is limited to 5 days because of concerns about overgrowth of bacteria that might have inadvertently contaminated the preparation.
The clinical response to platelet transfusion can be assessed by measuring the increment in platelet concentration achieved in the patient’s blood. This generally correlates directly with the dose of platelets infused and inversely with the patient’s size. Using physiologic principles, one can calculate what this response should be. Although the ideal theoretical response is occasionally achieved, on average, the response is approximately one-half of what one would predict because of immunologic and nonimmunologic clinical factors that impact negatively on the response. There is no single correct dose of platelets for all patients. On average, both the initial increment and the time to next transfusion will increase with increasing platelet dose. The appropriate dose will vary with the clinical circumstances and the patient’s size and individual response to transfusion. The traditional platelet concentration that should trigger a platelet transfusion had been 20,000/µl, but studies have shown that this level can safely be reduced to 10,000/µl in stable patients. It is important to raise the transfusion trigger above this level in response to a variety of clinical circumstances that increase the likelihood of bleeding. Although most platelet transfusions are given to patients whose marrows are suppressed, occasionally platelet transfusion is indicated when the thrombocytopenia is due to massive blood loss, cardiopulmonary bypass, splenomegaly, immune-mediated thrombocytopenia, and hereditary thrombocytopenia.
The complications of platelet transfusion are almost always due to contaminating leukocytes, red cells, plasma proteins, and microorganisms. Those due to contaminating leukocytes can be reduced in frequency by prestorage leukoreduction of the platelet products. Alloimmunization to class I HLA antigens can be managed by a variety of strategies using apheresis platelet concentrates that lack the antigens to which the patient has formed antibody.
Acronyms and abbreviations that appear in this chapter include: ACE, acetylcholinesterase; BSA, body surface area; CCI, corrected count increment; CMV, cytomegalovirus; CREGs, cross-reactive groups; DMSO, dimethyl sulfoxide; FDA, Food and Drug Administration; FNHTR, febrile nonhemolytic transfusion reactions; GMP, good manufacturing practices; GVHD, graft-versus-host disease; HIV, human immunodeficiency virus; HLA, human leukocyte antigen; ITP, immune (idiopathic) thrombocytopenic purpura; LCT, lymphocytotoxicity; PRA, percent reactive antibody; RhoGAM, Rh immunoglobulin; TTP, thrombotic thrombocytopenic purpura.
In the 1980s and early 1990s there was a rapid increase in the use of platelet transfusion with a doubling in the United States between 1982 and 19891,2 (Fig. 142-1). Platelet use in the United States peaked in 1992 at 9,330,000 units with a slight decline to 7,840,000 in 1994.3 In 1994, for the first time, platelets obtained by apheresis represented more than 50 percent of the platelets infused. The progressive increase in platelet use through the 1980s and early 1990s coincided with increasingly aggressive myelosuppressive therapy for malignancies and increased availability of platelets made possible by the development of cost-effective methods for storage of platelet concentrates.
FIGURE 142-1 Trends in platelet utilization in millions of whole-blood-derived platelet concentrate equivalent units per year. The data for platelet-rich plasma platelet concentrates and apheresis platelet concentrates are from the United States as reported in references 1,2 and 3. In 1994, for the first time, more than half of platelet transfusions were given as apheresis platelet concentrates. Also in 1994, a small but significant fraction were leukoreduced prior to storage. Between 1988 and 1994, an increasing number of platelet concentrate transfusions were prepared by the buffy coat method in Europe, incidence estimated from reference 135
TECHNIQUES FOR PLATELET PREPARATION
Platelet concentrates for transfusion may be obtained from routine donations of whole blood anticoagulated with citrate-based formulations or by apheresis with a variety of apheresis devices which also use citrate as the anticoagulant. Two methods of preparing platelet concentrates from whole blood are used, the platelet-rich plasma method and the buffy coat method.
In addition to appropriate platelet content, much attention is now being given to the level of contaminating leukocytes that are infused into patients. Problems produced by contaminating leukocytes will be discussed in the section on complications of platelet transfusion. In toto, these complications are sufficiently serious that many now recommend a totally leukoreduced blood supply. Blood products can be filtered during infusion at the bedside, but, for reasons to be discussed, it is probably preferable to accomplish leukoreduction at the time of preparation of the product. In the United States, an apheresis platelet concentrate or a pool of platelet-rich plasma platelet concentrates is considered leukoreduced if it contains less than 5 × 106 leukocytes. The standard in Europe is 1 × 106 leukocytes.
WHOLE-BLOOD-DERIVED PLATELET CONCENTRATES
Whole-blood-derived platelet concentrates are often termed random-donor platelet concentrates. This was to distinguish them from apheresis platelet concentrates derived from specific donors for specific refractory patients generally on the basis of HLA matching. Now, apheresis platelet concentrates are commonly given “randomly” to patients who do not require products from specific donors. Therefore, the term whole-blood-derived platelet concentrates is preferred.
PLATELET-RICH PLASMA PLATELET CONCENTRATES
This is the only method used in North America for whole-blood-derived platelet concentrates: 450 to 500 ml (a unit) of whole blood is held for up to 8 h at room temperature, and platelet-rich plasma is separated from red cells and buffy coat by low-speed centrifugation. After transferring the platelet-rich plasma to another bag, the plasma is centrifuged rapidly to produce a platelet pellet. Most of the plasma is removed, and then the platelet pellet is allowed to “rest” for 1 to 2 h before resuspension in approximately 50 ml of autologous citrated plasma. Without this rest period, platelets tend to clump irreversibly upon resuspension. The separated red cells are used for transfusion, while the supernatant plasma is used for transfusion or fractionation.
Over the past 20 years, technical improvements have resulted in an increase in the number of platelets in each unit from 5 to 6 × 1010 in 1975 to as much as 9 to 10 × 1010 in 1997.4 With modern techniques an average unit should contain approximately 8 × 1010 platelets. Approximately 1 to 5 × 108 leukocytes, predominantly lymphocytes, will also be present. One unit is adequate only for the transfusion of a small child less than 30 lb in weight. To transfuse adults, 4 to 8 units need to be pooled to provide a therapeutic dose (see section on dose below).
Whole-blood-derived platelet pools of 4 to 9 units have a high level of leukocyte contamination, 0.4 to 4.0 × 109, which is three orders of magnitude higher than that of a leukoreduced transfusion. There is a system which inserts a leukocyte-reduction filter between the primary blood bag and the bag that accepts the platelet-rich plasma.5 Thus the platelet-rich plasma is leukoreduced at the time of its preparation. This system was introduced in early 1998 for the preparation of all platelet-rich plasma platelet concentrates in Canada.
BUFFY COAT PLATELET CONCENTRATES
The buffy coat method is being used with increased frequency, particularly in Europe.6 An initial hard centrifugation sediments all blood cells so that the plasma, buffy coat, and red cells can be collected in three separate containers. Remarkably, not only do the platelets at the top of the bag fall to the buffy coat, but also the platelets at the bottom of the bag rise to the buffy coat. Therefore, platelet yields in the buffy coat are excellent. One can prepare platelet concentrates from individual buffy coats7 or pool 4 to 6 buffy coats, add 2 to 4 volumes of an additive solution, centrifuge the pool at low speed to remove red cells and leukocytes, and push the supernatant through a leukoreduction filter to produce a therapeutic, leukoreduced, dose of platelets for an adult.8,9
The platelet-rich plasma and buffy coat methods each have their advantages and disadvantages.6,10,11 Each produces platelets of high quality, and platelet yields are equivalent. In the buffy coat method, 20 to 25 ml of red cells are lost with the buffy coat, but an extra 70 to 80 ml of plasma can be collected. In any event, the buffy coat method is being used with increasing frequency in Europe; it is not used in North America.
APHERESIS PLATELET CONCENTRATES
One can obtain 2.5 to 10 × 1011 platelets (equivalent to 3 to 10 units of whole-blood-derived platelet concentrates) by apheresis of donors over 1 to 2 h using a variety of devices,12,13,14 and 15 with an extraordinarily high level of safety for the donor.16 The number of platelets obtained during the procedure varies according to the platelet concentration in the blood of the donor, the duration of the donation, and the efficiency of the device. The efficiency of the newest devices is such that one should expect to obtain at least 60 percent of the platelets that pass through them, and most donors begin to be quite restless if the donation time exceeds more than 90 to 120 min. Therefore, it is the wide range of the platelet concentration in the blood of normal donors, 130,000 to 500,000/µl that accounts for the wide range in platelet yields.17 There are also devices for obtaining both apheresis platelet concentrates and a unit of red cells from the same donor at the same sitting.18
The goal of apheresis is to obtain a therapeutic dose of platelets for an adult from a single donor during one apheresis sitting. In a subsequent section, we will discuss current controversies concerning the appropriate dose for platelet transfusion. Current standards of the FDA in the United States state only that 75 percent of apheresis platelet products must contain more than 3.0 × 1011 platelets. While 2.5 to 3.5 × 1011 is probably a satisfactory dose for the prophylactic transfusion of a child or small adult, it is probably unsatisfactory for a large adult who is bleeding or has other clinical features that interfere with an optimal response to platelet transfusion. On the other hand, administration of high-yield platelet products to small adults and children may be wasteful. Blood centers are in the process of considering the best way to handle the manufacturing process for apheresis platelets. Consideration is being given to the preparation of products containing two or more levels of platelet content, perhaps means of 3.2 × 1011 and 6.4 × 1011 (i.e., approximately 4 and 8 whole-blood-derived units respectively). The use of the products could be tailored to the needs of individual patients.
There are two major advantages in using apheresis platelet concentrates. First, the number of donors to whom the patient is exposed is substantially reduced, thus reducing the likelihood of transmission of viral and bacterial diseases. Second, the separation technology of the various apheresis devices lends itself to the production of leukoreduced products during collection. Progressive improvements of those most recently available12,13 has allowed the production of products with less than 1 × 106 leukocytes essentially 100 percent of the time. Figure 142-2 shows the experience of one blood center in this regard.
FIGURE 142-2 Cumulative frequency of leukocyte contents of apheresis platelet concentrates produced with six different apheresis methodologies: Fenwal Amicus, COBE Spectra (standard and leukocyte reduction system [LRS] versions), Fenwal CS-3000 (single- and dual-needle access), and Haemonetics MCS+. The newest devices, the Amicus and Spectra LRS, produce >99% products with leukocyte content less than 5 × 106, the current standard in the United States. The older procedures have a 25% (standard Spectra, dual-needle CS-3000) or 95% (single-needle CS-3000, MCS+) failure rate. Data from the American Red Cross Blood Services, Penn-Jersey Region. Reprinted from reference 136 with permission.
Prestorage leukoreduction at the blood center offers the advantage that it is carried out under standardized conditions following GMP with appropriate quality control procedures in place. Such standardization is not possible with filtration at the bedside.19 There are reports in which bedside filtration has failed to achieve the expected beneficial results.20
STORAGE OF PLATELET CONCENTRATES
LIQUID STORAGE AT 20 TO 24°C
Both whole-blood-derived and apheresis platelet concentrates may be stored for 5 days using the same principles: (1) The temperature must be 20 to 24°C.21 (2) The storage container must be constructed of a plastic material that allows adequate diffusion of oxygen to meet the cells’ metabolic needs.22,23 (3) The platelet concentrates must be agitated during storage.22,23
Using radiolabeling of stored platelets (Fig. 142-3), survival after reinfusion in vivo is nearly normal if storage, even for several days, is carried out at 20 to 24°C (68.0 to 75.2°F). However, at colder temperatures, survival is dramatically shortened.21 If oxygen influx is inadequate at 20 to 24°C (68.0 to 75.2°F), the cells will increase their production of lactic acid, leading to depletion of bicarbonate buffer and fall in pH to less than 6.2.22,23 These acid conditions result in the platelets being rapidly cleared from the circulation after transfusion. A similar fall in pH occurs if the platelet concentrates are not agitated during storage.23 There are data suggesting that agitation may be discontinued for up to 24 h of the 5-day storage interval.24
FIGURE 142-3 Relationship between storage temperature and platelet viability after transfusion. Platelet-rich plasma was obtained from normal volunteers and stored overnight at the indicated temperatures. Thereafter, the platelets were labeled with radioactive chromium and reinfused. Percent yield refers to the percent of platelets infused that circulate in the first 3 h after infusion. The 50–60% yield at 22°C (71.6°F) is a result of physiologic pooling in the spleen (see Chap. 117), not cell damage. The combination of percent yield and subsequent in vivo survival (T1/2) is optimal at 22°C.
Synthetic media are also used for the storage of buffy coat platelet concentrates.8 Definition of the optimal solution is still in progress, but it appears that it can be relatively simple, relying on acetate as an oxidative fuel for platelets.25 The oxidation of an organic anion such as acetate utilizes a proton from the medium, thus providing an alkalinizing effect that spares bicarbonate, the major buffer during platelet concentrate storage.26 It is anticipated that synthetic media will soon be used for apheresis platelet concentrates as well.27
Some investigators have been unable to find any practical difference in clinical response between fresh and stored platelets,28,29 but most find a reduction in recovery in vivo, with survival reduced by approximately 20 to 25 percent after 5 days of storage as judged by radiolabeling studies in normal volunteers and by the increase in platelet count in thrombocytopenic patients.30 Furthermore, some authors have reported an even greater defect in stored platelets relative to fresh platelets in sick patients with fever, sepsis, splenomegaly, and disseminated intravascular coagulation.31,32
Platelet recovery and survival is as satisfactory after 7 days of storage as it is after 5 days.33 However, when storage was extended to 7 days, bacterial overgrowth and clinical sepsis in recipients of stored platelets occurred with sufficient frequency to warrant limiting liquid storage to 5 days.34 If methods of bacterial decontamination35 or bacterial detection are developed,36 it may be possible to prolong storage beyond 5 days once again.
Many platelet abnormalities have been described after ex vivo platelet storage.37 At present, in vitro characteristics that correlate best with the capacity to circulate in vivo are retention of disc shape and good function in the hypotonic shock response.38 With few exceptions, platelets with normal discoid morphology will circulate normally after transfusion. Platelets that are damaged by cold, acidity, or bacterial contamination generally lose their discoid morphology and become spheres. Normal discoid morphology is reflected by the “swirling” or “shimmering” appearance of well-preserved platelet concentrates during gross, visual inspection.39 Blood bank staff and clinical personnel are urged to check platelet concentrates for this phenomenon prior to transfusion (Fig. 142-4).
FIGURE 142-4 Swirling of platelet concentrates. Platelets in platelet concentrates that have been prepared and stored well retain their normal discoid configuration, which confers a swirling or shimmering appearance to the platelet concentrates (left). If the platelets are damaged by cold temperature, a fall in pH, or bacterial contamination, discoid shape and the swirling appearance are lost (right). Loss of swirling allows a transfusion unit or a clinician to identify potentially ineffective or dangerous platelet concentrates.
The activities of coagulation factors are well maintained in the suspending plasma of platelet concentrates during storage, except for modest decreases in those of factors V and VIII.40 Thus, a pool of 4 to 8 whole-blood-derived platelet concentrates or an apheresis platelet concentrate provides the equivalent of 1 to 2 units of fresh-frozen plasma.
The most widely used method for frozen storage employs controlled rate freezing (1°C per minute), 5% DMSO as a cryoprotective agent, rapid thawing, graded reduction of the DMSO concentration, and washing prior to infusion. In vivo viability is approximately 40 to 50 percent relative to fresh platelets.41 Thus, this technology is both more complex and expensive and less effective than liquid storage at 20 to 24°C.42 However, these preparations can be effective clinically43 and may be very valuable for autologous transfusion of selected patients who may not respond well to allogeneic platelets. Platelets may be obtained before myelosuppressive therapy, then frozen and administered during subsequent periods of thrombocytopenia.44 Newer approaches using second-messenger effectors may allow the use of lower concentrations of DMSO, which would, in turn, allow the direct infusion of platelets after thawing.45
FRESH WHOLE BLOOD
Children under the age of 2 who undergo complex cardiac bypass surgery have a better hemostatic response to fresh whole blood than to reconstituted blood using red cells, plasma, and platelet concentrates prepared as described above.46 These are the only published data suggesting that the process of platelet concentrate preparation impairs platelet function. Although this conclusion has not been confirmed in a second trial, the administration of fresh whole blood is strongly endorsed by some cardiac surgeons.
LYOPHILIZED PLATELETS, PLATELET MEMBRANES, AND PLATELET SUBSTITUTES
Current methods of platelet storage are cumbersome, and the duration of storage is short. It would be ideal to have a safe and effective dried preparation with long shelf-life that one could simply rehydrate and infuse. A great deal of research is going on in this area examining platelets treated with paraformaldehyde and lyophilized platelet membrane microvesicles, fibrinogen-coated beads, albumin microspheres, and others.47 This is an important area, but all these ideas await validation in appropriate clinical trials.
There are also nontransfusional drugs, including the antifibrinolytic agents aminocaproic acid and tranexamic acid, that may help to stop thrombocytopenic bleeding.48 They have been described as being effective in controlling mucosal and dental bleeding in thrombocytopenic patients without increasing the platelet concentration in the blood.
GENERAL PRINCIPLES IN PATIENTS WITH MARROW FAILURE
Assuming that one-third of infused platelets will be pooled reversibly in a spleen of normal size (see Chap. 117) and that the recipient’s blood volume is 2.5 liters/m2, the infusion of 1 unit of whole-blood-derived platelet concentrates, containing 8.0 × 1010 platelets into a recipient with 1 square meter of BSA, should result in an increase in platelet count of 21,000/µl. Of course, the response to 1 unit will be inversely proportional to the patient’s size, expressed as the body surface area. Thus, one can evaluate the response to a platelet transfusion by calculating the corrected count increment, or CCI49:
The measurement of CCI has its critics,50 but it is the most widely used method. Under optimal circumstances, the response should be 21,000/µl per square meter per unit infused or 26,000/µl per square meter per 1011 platelets infused.
In practice, in patients with thrombocytopenia secondary to marrow failure, the average CCI is approximately one-half of that expected, 10,000/µl per square meter per unit infused (Fig. 142-5).49 Many studies have attempted to identify the factors responsible for this consistent but less than optimal response.32,49,51,52,53,54 and 55 Alloimmunization has been incriminated, along with a variety of nonimmune factors such as platelet storage, bacterial sepsis, concomitant use of antibacterial antibiotics and amphotericin B, graft-versus-host disease, splenomegaly, disseminated intravascular coagulation, and simply having had a recent allogeneic bone marrow transplantation. It is of interest that no one factor predominates in the majority of studies, suggesting that the crucial factors vary with the populations of patients being studied.
FIGURE 142-5 Rise in platelet concentration 1 h after infusion in patients with acute leukemia (redrawn from reference 49). As described in text, the increment in concentration for each unit infused has been corrected for body surface area. Without complicating factors, it should be approximately 21,000/µl (21 × 109/liter). The vertical axis refers to the percent of transfusions achieving the increments indicated. There is marked heterogeneity in response, with a median of approximately 10,000/µl (10 × 109/liter), approximately one-half of what one would predict.
The initial CCI can be measured in 10 min56 to a few hours after the transfusion. The time until the platelet concentration returns to baseline (time to next transfusion) also varies with the immune and nonimmune factors affecting the initial CCI.57 It also varies directly with the height of the platelet count achieved by the transfusion and, therefore, the dose of platelets administered.58,59 and 60 This follows from the fact that platelet survival is reduced in all patients with thrombocytopenia, regardless of the cause, with progressive reduction as the platelet concentration in the blood becomes lower61 (see Chap. 117). Therefore, everything else being equal, the larger the dose, the higher the platelet count achieved by transfusion, and the longer the time will be to the next transfusion (Fig. 142-6).
FIGURE 142-6 Response to varying doses of units of whole-blood-derived platelet concentrates during repetitive transfusion of patients thrombocytopenic from marrow failure. Larger doses produce not only an initial higher increase in platelet concentration but a longer time to next transfusion. Fewer transfusion episodes are required over a 20-day transfusion period. Graph redrawn with permission from calculations in reference 58, validated by empirical data given in references 59 and 60.
If the average patient has an increase in platelet concentration of 10,000/µl per square meter per unit of platelets, one can calculate the relationship between the dose administered and the platelet count achieved. Furthermore, using calculations and data from references 58,59 and 60, one can estimate the time to the next transfusion. For example, for the transfusion of a myelosuppressed patient of a 2-m2 surface area and platelet count 5000/µl, these values are given in Table 142-1. The results indicate that there is no standard dose that fits all patients, regardless of size and clinical situation. For the doses given, the platelet concentrations achieved in a patient with 1-m2 body surface would be twice those in the table. In the 1-m2 patient, 4 units (3.2 × 1011) would probably be satisfactory if the patient is not bleeding, is being transfused prophylactically, and is hospitalized so that he or she can be transfused again 24 to 48 h later, since raising the platelet concentration to 20,000 to 30,000/µl protects most thrombocytopenic patients against spontaneous, catastrophic bleeding. However, 10 units (8.0 × 1011) would be a better choice if the goal was to achieve a platelet concentration over 50,000/µl because the patient was bleeding or was being prepared for an invasive procedure.62 It is probably most critical to achieve this level of platelet concentration if the surgical field is highly vascular, as with inflammation or portal hypertension; if there are coexisting defects in plasma coagulation; if the procedure is “blind,” as in needle biopsy of the liver, when there is no opportunity to achieve hemostasis mechanically; or if the surgery is in an area where even a small hemorrhage could be disastrous, as in the central nervous system.
TABLE 142-1 RELATIONSHIP BETWEEN PLATELET DOSE AND CLINICAL RESPONSE IN A PATIENT WITH A 2-M2 BODY SURFACE WITH A PLATELET COUNT, 5000/µl, PRIOR TO TRANSFUSION
A higher dose might also be chosen to facilitate transfusion in the outpatient setting where a longer transfusion interval would be preferable.
However, patients vary dramatically in their responses. Therefore, the measurement of increments 1 and 24 h after transfusion is a cost-effective way to modify the dose and frequency of transfusion based on the pathophysiology of the individual patient.
PLATELET TRANSFUSION TRIGGER
Clearly, a thrombocytopenic patient who is actively bleeding requires platelet transfusion. It is more difficult to make this judgment if the platelet count is simply very low and the patient has no hemorrhagic signs or only minor ones, such as petechiae or small ecchymoses of the skin. Clinical experience suggests that if the platelet count is low enough, long enough, “spontaneous” major hemorrhage, particularly into the central nervous system, may occur. Unfortunately, in an individual patient, we do not know how low and how long. It would be very helpful if a laboratory test were available that could be used to supplement measuring the platelet concentration in predicting bleeding.63
Studies of patients with acute leukemia carried out 35 years ago, prior to the availability of platelet transfusion, described the relationship between platelet concentration in the blood and clinical hemorrhage.64 Minor and major hemorrhage began when the platelet count fell below 50,000 and 20,000/µl respectively. Major hemorrhage was observed in the range 5000 to 20,000/µl but on only 3 percent of patient days. There was a rapid increase in the rate of major bleeding when the platelet concentration fell below 5000/µl, reaching a frequency of 33 percent of patient days as the platelet concentration approached 0/µl. Many of these children were, however, undoubtedly being treated with aspirin for pain and fever. Therefore, these bleeding rates may be overestimates.
Subsequently, the same group described the effect of prophylactic platelet transfusion administered whenever the platelet count fell below 20,000/µl.65 Although there was a striking reduction in major hemorrhage when the platelet count (measured pretransfusion) was below 5000/µl, there was no substantial change when the platelet count was in the range of 5000 to 20,000/µl. Nonetheless, for many years, this experience was used to justify prophylactic platelet transfusion whenever the platelet concentration fell below 20,000/µl, although the data actually suggested 5,000/µl as an appropriate trigger.
More recently, prospective but uncontrolled studies by one group supported the safety and efficacy of a more restrictive policy using 5000/µl as the platelet transfusion trigger.66,67 Subsequently, three prospective studies assigned patients to one of two groups receiving prophylactic platelet transfusion at either 10,000/µl or 20,000/µl.68,69 and 70 Uniformly, there was no increase in bleeding risk at the lower transfusion trigger, which is now being adopted by many transfusion services.71
However, it is just as unjustified to choose a rigid transfusion trigger as it is to choose a rigid transfusion dose. It is commonly assumed that clinical factors increase the risk of hemorrhage at any given platelet concentration. These include fever and sepsis, administration of drugs that interfere with platelet function, coexistent abnormalities of plasma coagulation factors, disseminated intravascular coagulation, and high leukocyte concentrations in the blood. Thus, it is appropriate to raise the transfusion trigger in complicated, clinically ill patients.
In addition, moderate to severe bleeding is observed in 11 to 23 percent of patients after marrow transplantation in spite of the aggressive use of prophylactic platelet transfusion and maintenance of morning platelet counts above 20,000/µl.72,73 Gastrointestinal and urinary bleeding are most common; pulmonary and intracranial bleeding are less common. Usually, there is an identifiable anatomic cause, such as gastrointestinal ulceration, hemorrhagic cystitis, or diffuse alveolar hemorrhage. In effect, it is common in the myelosuppressed patient to be treating bleeding, not preventing it. Many centers increase the transfusion trigger to 30,000 to 50,000/µl in such cases.72
THROMBOCYTOPENIA DUE TO PLATELET LOSS, SEQUESTRATION, OR DESTRUCTION
Dilutional thrombocytopenia will occur when massive blood loss is replaced with units of stored red cells that lack viable platelets. Following replacement of one blood volume, 35 to 40 percent of the platelets usually remain. Even when one to two blood volumes have been replaced, abnormal bleeding usually does not develop, and routine transfusion is not indicated simply because the platelet count is low.74 Platelets should be given to patients who demonstrate abnormal bleeding.
Immediately following and for several days after cardiac surgery, the platelet count commonly falls to subnormal levels, occasionally as low as 50,000/µl (50 × 109/liter). There is an associated defect in platelet function. Prospective studies have shown no benefit from the prophylactic administration of platelet transfusions to such patients.75 They should be reserved for the relatively rare patient who demonstrates clinically abnormal bleeding.
Patients with massive splenomegaly have thrombocytopenia related predominantly to excessive sequestration in a splenic pool in continuous exchange with platelets in the circulation (see Chap. 117). The platelet count rarely falls below 30,000/µl (30 × 109/liter) due to this mechanism alone, so that platelet transfusion is rarely considered except in anticipation of invasive procedures such as surgery and needle biopsy of the liver. Under these circumstances, depending on the degree of splenic enlargement, one may need to administer 10 to 15 units of whole-blood-derived platelet concentrates per square meter body surface area to achieve a substantial increase in platelet count. In many patients with severe splenomegaly, it may not be possible to achieve platelet count elevations even with large numbers of platelet concentrates. If such patients require elective surgery, consideration should be given to splenectomy prior to the elective procedure.
IMMUNE (IDIOPATHIC) THROMBOCYTOPENIC PURPURA (ITP)
In ITP, platelet transfusion is generally not used because the bleeding tendency is less severe than in thrombocytopenia due to diminished production, and the response to medical therapy is generally satisfactory and rapid (see Chap. 117). Furthermore, the survival of transfused platelets is relatively brief, similar to that of the patient’s own platelets. Nonetheless, when there is critical bleeding or need for urgent surgery, 3 to 6 units of whole-blood-derived platelet concentrates per square meter body surface area will generally raise the platelet count for 12 to 48 h.76 The same general principles apply to other diseases in which there is accelerated destruction of platelets, such as disseminated intravascular coagulation.
NEONATAL ALLOIMMUNE THROMBOCYTOPENIA
In this syndrome, the mother produces an alloantibody against antigens on fetal platelets that have crossed the placenta (Chap. 117). The antibody, in turn, crosses the placenta, causing in utero thrombocytopenia that may persist for weeks after delivery. Since maternal platelets are compatible, platelets harvested from the mother’s blood by apheresis can produce an adequate increase in the infant’s platelet count after infusion.77 Ideally, such platelets should be concentrated in a small volume of plasma or washed, so as to avoid infusing additional antibody.
Unfortunately, it is often difficult to arrange for apheresis of the mother. Surprisingly, a randomly selected unit of platelets may raise the neonate’s platelet concentration substantially.78 If it does not, prompt serologic evaluation of the mother and father can identify the antigen to which the antibody has been formed, usually HPA-1a (PlA1). Donors lacking HPA-1a (i.e., homozygous HPA-1b) can then be recruited to support the infant.79
These syndromes are rare and generally not associated with severe bleeding.80 Since the survival of allogeneic platelets is normal, platelet transfusion is quite effective and may be used for critical bleeding and surgery.
QUALITATIVE PLATELET DISORDERS
In spite of normal platelet counts, patients with qualitative platelet disorders have a clinical bleeding tendency associated with abnormal in vitro tests of platelet function and a prolonged bleeding time in vivo. The basis may be hereditary (see Chap. 119) or acquired (see Chap. 120). Platelet transfusion is generally not indicated when the cause is extrinsic to the platelet, as in uremia, von Willebrand disease, and hyperglobulinemia, since the transfused platelets will function no better than the patient’s own platelets. There are exceptions in certain types of von Willebrand’s disease in which normal platelets can be used to deliver von Willebrand factor to a bleeding site (see Chap. 135). Most inherited intrinsic disorders are mild and do not require platelet transfusions even for surgery if the procedure is carried out under direct vision, so that hemostasis may be achieved mechanically. If the bleeding tendency is more severe, as in Glanzmann thrombasthenia, platelet transfusions may be necessary for more severe bleeding and surgery. The acquired defects, as in the myeloproliferative and myelodysplastic syndromes, generally do not require platelet transfusion unless there is coexistent thrombocytopenia.
POSSIBLE CONTRAINDICATIONS TO PLATELET TRANSFUSION
Concern has been voiced that platelet transfusions should not be administered to patients with forms of thrombocytopenia associated with thrombosis such as TTP and heparin-induced thrombocytopenia (see Chap. 117) since infusion of platelets might worsen the thrombotic tendency.81,82 Unfortunately, particularly in TTP, platelet transfusion is often requested prior to invasive procedures such as the insertion of intravenous catheters for therapy with apheresis. The author’s experience has been that platelet transfusions are safe in this setting. However, it seems prudent not to administer prophylactic platelet transfusions simply because the platelet concentration is low in these diseases.
COMPLICATIONS OF PLATELET TRANSFUSION
There are many complications of platelet transfusion (Table 142-2). Paradoxically, they are not due to the platelets but rather to contaminating leukocytes, red cells, plasma proteins, and microorganisms.
TABLE 142-2 COMPLICATIONS OF PLATELET TRANSFUSION
COMPLICATIONS DUE TO CONTAMINATING LEUKOCYTES
ALLOIMMUNIZATION TO CLASS I HLA ANTIGENS
HLA antigens are expressed on integral membrane glycoproteins. Almost all cells have Class I antigens (A, B, and C subloci), whereas only a few types of circulating leukocytes (dendritic cells, monocytes, and subsets of B cells) have Class II antigens. Primary alloimmunization to Class I HLA antigens appears to require presentation of such antigens on cells that also express Class II antigens and other costimulatory molecules.83 There is now abundant evidence that the incidence of HLA alloimmunization can be reduced by the consistent use of leukoreduced blood products.84,85 Transfused red cells must also be leukoreduced, since leukocytes contained in them are quite capable of inducing HLA alloimmunization.86
However, it remains to be seen how completely leukoreduction can eliminate alloimmunization. In one study,87 leukoreduction had almost no effect on the development of alloimmunization in patients who had been previously exposed to foreign leukocytes either through pregnancy or transfusions that had not been leukoreduced. It may be that leukoreduction will prove to be less effective in preventing secondary as opposed to primary alloimmunization. However, another trial84 showed efficacy in such patients.
Alloimmunization to HLA should be suspected clinically if two or three consecutive platelet transfusions produce a CCI less than 3000/µl per square meter per unit.88 It can be confirmed in the laboratory by performing a lymphocytotoxicity (LCT) assay for HLA antibody in the patient’s serum. In this assay, leukocytes from 50 to 100 donors with an appropriate heterogeneity of HLA types are incubated with the patient’s serum and complement. LCT is assessed microscopically. The presence of such antibody has been a good predictor of poor response to platelets from randomly selected donors89 and improved response when platelets are matched for HLA type.90 Many centers consider it inappropriate to issue matched platelets unless this, or a similar test, has been performed because of the belief that matching will provide little benefit if antibody is not present.91,92
The incidence and severity of HLA alloimmunization should be gradually decreasing as the use of leukoreduced blood products becomes increasingly popular. The following discussion is based on data accumulated before widespread leukoreduction was in use. Approximately 10 percent of patients presenting for therapy of diseases requiring platelet transfusion will already have LCT antibodies from prior transfusions and pregnancies.93 Another 30 percent become alloimmunized during therapy, and 60 percent never do.93 There is no known difference between those who do and do not become immunized. Among those who do, some will do so after only two to four transfusions, whereas others require dozens of transfusions.94 The pattern and intensity of immunization varies greatly from patient to patient. An LCT assay can be characterized by the percentage of cells in the panel against which the patient’s serum reacts, i.e., the percent reactive antibody (PRA). Furthermore, the pattern of reactivity can be analyzed to determine the antigens to which the patient has formed antibody. Patients may have PRA values between 4 and 100 percent (Fig. 142-7).
FIGURE 142-7 Cumulative frequency of percent reactive antibody (PRA) by lymphocytotoxicity (LCT) assay in serum samples from 108 thrombocytopenic patients referred to a regional blood center for matched platelet support during a 6-month period. Fifty showed no reactivity, while 58 showed PRAs that varied from 4% to 100%. The squares indicate patients who had demonstrable antibody against antigens within their own cross-reactive groups, i.e., intra-CREG antibody. Data provided by Dr. Susan Hsu, American Red Cross Blood Services, Penn-Jersey Region. Reprinted from reference 136 with permission.
The majority of patients who become alloimmunized establish a level and specificity of immunization and tend to maintain that status as they continue to be transfused. However, approximately 30 percent lose their antibodies over time in spite of continuing transfusion.95 Thus, it is helpful to monitor antibody levels and specificity, since such patients may regain responsiveness having been previously refractory.
Cross-reactive groups (CREGs) of HLA antigens have been defined by serologic testing. Cross-reactivity among antigens in a CREG is based on the sharing of one or more public epitopes by those antigens.96 It is common for some patients to develop antibodies to one or more public epitopes, i.e., one or more CREGs, while others develop antibodies to one or more private antigens.97 Furthermore, some patients demonstrate intra-CREG antibodies, i.e., antibodies to antigens in the same CREG as the patient’s own antigens.98 These facts have major implications for management.
MANAGEMENT OF THE REFRACTORY PATIENT
In 1969, it was shown that refractory patients would respond to platelets from siblings who were identical for all four Class I HLA antigens.99 In fact, this simple clinical observation remains one of the most compelling pieces of evidence supporting the role of HLA alloimmunization in platelet refractoriness. Similarly, patients could be supported by platelets from unrelated donors who were HLA-identical or closely matched. Because some patients do not make antibody to antigens within their own CREGs, it became popular to choose donors according to CREG classification, particularly BX matches, i.e., donors whose antigens are identical to or within the same CREGs as those of the patient.100 Table 142-3 shows the categorization of such matches.
TABLE 142-3 CLASSIFICATION OF DONOR/RECIPIENT PAIRS ON THE BASIS OF HLA CLASS I MATCHING
Responses were better with such matching than with random donor selection, but many BX matches failed and many C and D matches succeeded. Figure 142-7 suggests potential explanations for the relatively poor predictive capacity of this method. Some patients with relatively low PRAs have antibody to only one or two CREGs so that success with some C and D matches would be expected. On the other hand, failure of some BX matches would be expected in patients with intra-CREG antibody. Futhermore, this method generally does not provide a good match quickly. It is uncommon to find an excellent match in a blood center’s inventory, and days are required to recruit and pherese one or more well-matched donors from an HLA-typed donor file.
In the late 1980s and early 1990s, practical methods for platelet cross-matching became available.101 Many centers found that they could simply cross-match patient’s serum with unselected apheresis platelet concentrates in inventory to find, within hours, a compatible product that would be successful in vivo.51,102 However, it was recognized102 that, in a highly immunized patient, one might cross-match with dozens of donors and not find a compatible product. For many such patients, only the identification of an A or BU match (Table 142-3) will suffice.
One other approach to these patients has been proposed.103,104 If the PRA in the LCT assay is less than 100 percent, one should be able to identify the HLA antigens to which the patient has not formed antibody. The patient can then be supported with “antigen-negative” platelets, i.e., platelets which lack the antigens to which the patient has formed antibody. If the results of the LCT assay are known, one can often provide a product from inventory on an urgent basis using this approach.
There is no reason that one of these approaches should be chosen to the exclusion of the other two.105 If the PRA is less than 70 percent, successful support can usually be provided by cross-matching of random products or by “antigen-negative” platelets. When the PRA is high (>80%), one can select for cross-matching the best available “antigen-negative” HLA matches and/or selectively recruit A and BU matches from an HLA-typed donor file. Some patients with common HLA types, 1,2/7,8 for example, will have dozens of A and BU matches available. Unfortunately, some patients have rare HLA types and will have no A or BU matches to recruit.
ROLE OF ABO AND PLATELET-SPECIFIC ANTIBODIES
ABO determinants are carried by both glycoproteins and glycolipids of platelets, as they are on red cells.106 Group O patients commonly have a good response to platelets from group A or B donors, but in a subset of patients, the response can be poor107 or very poor.108 Thus it seems wise to observe ABO compatibility when possible.
The platelet surface carries many platelet-specific antigens (HPA, see Chap. 138) which are quite capable of eliciting a strong alloantibody response. It is surprising that there are only a few case reports of such alloantibodies accounting for refractoriness to platelet transfusion.109,110,111 and 112 This area deserves further study, since it is technically difficult to identify platelet-specific antibodies when there is intense HLA alloimmunization as well. Techniques for doing so are improving.112,113
FEBRILE NONHEMOLYTIC TRANSFUSION REACTIONS (FNHTR)
Prior to the availability of methods for leukoreduction, approximately 20 percent of platelet transfusions were accompanied by FNHTR.114 Some of these reactions were undoubtedly due to antibodies in the patient directed against either leukocyte-specific or HLA antigens on leukocytes contaminating the platelet product. Leukocyte depletion by filtration during infusion reduced the frequency of these reactions, but many continued to occur.114,115 It is now clear that contaminating leukocytes produce inflammatory cytokines such as interleukin-1, interleukin-6, interleukin-8, and tumor necrosis factor alpha during storage at 20 to 24°C and that these compounds are responsible for many FNHTR since they are not removed by bedside filtration.116,117 These reactions provide a strong argument for routine, prestorage removal of leukocytes.
Nonetheless, FNHTR occur in approximately 2 percent of platelet transfusions even with prestorage leukoreduction.118 The cause of these reactions is not known. They may be related to plasma proteins or products produced during storage by the platelets themselves. In these rare cases, one can wash the platelets free of plasma prior to infusion.119
TRANSMISSION OF CYTOMEGALOVIRUS (CMV)
In asymptomatic carriers, this virus resides in the nuclei of subsets of leukocytes with little virus free in plasma. It has been shown that the use of leukoreduced blood components is essentially equivalent to the use of components from CMV-negative donors in terms of risk of CMV transmission.120 Since this infection is particularly dangerous for severely immunocompromised patients, such as those who have had a recent allogeneic bone marrow transplantation, some clinicians continue to use CMV-negative blood products along with leukoreduction in this select population.
GRAFT-VERSUS-HOST DISEASE (GVHD)
Immunosuppressed patients may develop GVHD from T lymphocytes present in any transfusion, including platelet transfusion. Thus, it is standard practice to treat platelet concentrates with gamma irradiation to inhibit proliferation of these T lymphocytes when the recipient had been heavily immunosuppressed.121 Clinicians are progressively applying this practice to all patients who have received cytotoxic chemotherapy. Exposure to 5000 rad appears to have no deleterious effect on platelets. It is important to emphasize that current methods of leukoreduction do not remove enough T cells to prevent GVHD.
COMPLICATIONS OF FILTRATION OF PLATELET CONCENTRATES
Finally, there may be complications of platelet transfusion related to the removal of leukocytes by filtration of platelet concentrates at the bedside. Severe hypotension has been reported,122 predominantly when negatively charged leukocyte reduction filters are used in patients who are receiving ACE inhibitors.123 One proposed mechanism is that high-molecular-weight kininogen is converted to bradykinin, a potent vasodilator, by exposure to a negatively charged surface. Bradykinin is normally metabolized by ACE in a few seconds but may circulate much longer in patients receiving ACE inhibitors. Some investigators have found no clinically significant contact system activation by these filters.124 Thus, the mechanisms behind these hypotensive reactions are still hypothetical, and more work needs to be done.
COMPLICATIONS DUE TO CONTAMINATING RED CELLS
When transfusing platelet concentrates to women of child-bearing potential who are Rh-negative, one needs to be concerned about sensitization by Rh-positive red cells contaminating infused platelets. In practice, sensitization is rather uncommon in immunosuppressed patients.125 However, where possible, one should administer platelets from Rh-negative donors. When this is not possible, one can administer Rh immunoglobulin (RhoGAM), about 20 µg intramuscularly per unit of platelets, so that the infused red cells will be cleared prior to sensitization. A full dose of RhoGAM, 300 µg, is sufficient to suppress the immune response to 15 ml of Rh-positive red cells.
There are enough red cells contaminating platelet concentrates to transmit both malaria and babesiosis if the donor is parasitemic with these infections.
COMPLICATIONS DUE TO PLASMA AND ITS CONTENTS
Storage of platelet concentrates at 20 to 24°C allows proliferation to dangerous levels of bacteria that occasionally contaminate units of blood or apheresis platelet concentrates.34 Contamination may occur because of asymptomatic bacteremia in the donor at the time of venipuncture, inadequate decontamination of the skin, or because of venipuncture through areas of the skin where bacterial colonization is deeper than can be reached by such decontamination.126 Bacterial contamination that might not be clinically significant after 2 to 3 days of storage may become so after 5 to 7 days.127 As previously mentioned, platelet concentrates storage is limited to 5 days for this reason.
The magnitude of this problem is commonly underestimated.128 Estimates suggest that there may be contamination of 5 to 10 units per 10,000 whole-blood-derived platelet concentrates and 150 clinical episodes associated with severe morbidity and death in the United States per year.129 This is 50- to 250-fold higher than the risk of mortality from transmission by transfusion of HIV or hepatitis B or C infection.
There are several potential approaches to this important problem. Because apheresis involves only one donor and one venipuncture, there may be less risk than for pooled whole-blood-derived platelet concentrates, although this has not been conclusively demonstrated. Under investigation are methods for screening platelet products for bacteria before transfusion36 and methods of viral inactivation which also inactivate bacteria.35 The methods of viral inactivation may inactivate T lymphocytes and prevent GVHD as well.130
The plasma diluent of platelet concentrates can transmit viruses such as hepatitis B and C and HIV. Recently improved methods of donor screening and testing have reduced but not eliminated this risk. As mentioned, methods for viral inactivation are being sought.35 Transmission of the parasite, Trypanosoma cruzi, which is responsible for Chagas disease, has also occurred with platelet transfusion.
Many transfusion services attempt to transfuse ABO-identical platelet concentrates. However, this is not always possible. When anti-A or anti-B is transfused to a patient whose red cells carry A or B, a positive direct antiglobulin test may be observed in the laboratory, making red cell compatibility testing more difficult.131 Actual accelerated destruction of the patient’s red cells is rare.132 However, on very rare occasions, frank acute hemolysis has been observed.133
As with any plasma infusion, one may encounter urticaria or anaphylactic shock in a patient with IgA deficiency and circulating anti-IgA, or transfusion-associated acute lung injury when a donor has leukocyte antibodies that can react with antigens on the leukocytes of the recipient.134
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Ernest Beutler, Marshall A. Lichtman, Barry S. Coller, Thomas J. Kipps, and Uri Seligsohn