Laurence Corash

Functional Characteristics of S-59 Photochemically Treated Platelet Concentrates Derived from Buffy Coats

Background: A photochemical treatment (PCT) process for inactivation of infectious pathogens and leukocytes has been developed and evaluated using single‐donor platelet concentrates. This study assessed the application of PCT to platelets prepared from pooled buffy coats. In this study, in vitro functional characteristics of PCT platelets were compared to control platelets prepared from pooled buffy coats using the approved platelet‐additive solution T‐Sol®. Platelets in platelet PAS III additive solution without PCT were evaluated as well. PCT also included the use of a psoralen (S‐59) reduction device (SRD). Materials and Methods: Four types of platelet concentrates were compared: (1) platelet concentrate in plasma/T‐Sol; (2) platelet concentrate in plasma/PAS III; (3) platelet concentrate in plasma/PAS III, PCT, 9 h SRD and (4) platelet concentrate in plasma/PAS III, PCT, 16 h SRD. PCT occurred on the day after whole‐blood collection. In vitro assay parameters included: pH, pO2, pCO2, HCO–3, platelet count, mean platelet volume, plasma glucose, plasma lactate, total ATP, expression of p‐selectin, hypotonic shock response and electron microscopy. Results: The results indicate that PCT is compatible with platelet concentrates prepared from pooled buffy coats for up to 7 days of storage. Conclusion: The PCT process resulted in acceptable in vitro platelet functional characteristics and is currently in clinical trials to evaluate the haemostatic efficacy of PCT platelets in thrombocytopenic patients requiring multiple platelet transfusions.

Corrected count increment and percent platelet recovery as measures of posttransfusion platelet response: problems and a solution.

BACKGROUND: Corrected count increment (CCI) and percent platelet recovery (PPR) are measures of response to platelet transfusion that “correct” the count increment for blood volume and number of platelets transfused. Their potential for data distortion is described, and a regression analysis is suggested that is more informative and avoids the inherent problems associated with using ratios as outcome measures.

PHOTOCHEMICAL DECONTAMINATION OF BLOOD COMPONENTS CONTAINING HEPATITIS B AND NON-A, NON-B VIRUS

Diluted plasma samples containing 10(2), 10(3), 10(4), and 10(5) chimpanzee infectious doses (CID) of a human non-A, non-B hepatitis virus (NANBV) were treated with a combination of two psoralen compounds, 4-aminomethyl-4,5,8-trimethylpsoralen and 4,5,8-trimethylpsoralen, and exposed to long wavelength ultraviolet. Each dilution was then transfused into one of four chimpanzees. In a second experiment, three samples containing 10(4.5) CID of hepatitis B virus (HBV) and two samples containing 10(4) CID of NANBV were treated with 8-methoxypsoralen (8-MOP) and ultraviolet irradiation. Two chimpanzees were each transfused with both a treated HBV and a treated NANBV sample. The third chimpanzee was inoculated with a treated HBV sample alone. In the six months after inoculation none of the animals showed biochemical or histological evidence of hepatitis. In experiments involving NANBV inocula, the susceptibility of the animals was confirmed by subsequent challenge with untreated NANBV. Factor VIII concentrate containing virus and photochemically treated as in the first experiment retained an average of 91% of its activity while that in the second experiment retained 94% of its activity.

The effect of a salmon diet on blood clotting, platelet aggregation and fatty acids in normal adult men.

This study was designed to measure the effect of dietary n−3 fatty acids (FA) on platelets and blood lipids. Healthy men (n=9), ages 31 to 65, were fed diets in which salmon was the source of n−3 fatty acids. They were confined in a nutrition suite at this Center for 100 days. Food intake and exercise levels were rigidly controlled. Initially they were placed on a stabilization diet for 20 days, then six men were fed the salmon diet for 40 days. The others remained on the stabilization diet. The two groups switched diets for the last 40 days of the study. Both diets were isocaloric [16% protein, 54% carbohydrate, and 30% fat by energy-% (En%)]. The salmon diet contained 7.5% of calories from n−6 FA and 2% from n−3 FA, primarily eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in a 40∶60 ratio, while the stabilization diet contained 7.5% of calories from n−6 FA and less than 0.3% n−3 FA, mainly 18∶3n−3. The bleeding time was unaffected by the diets in this study. The prothrombin time was shortened (11.6 sec.vs. 12.6 sec., p<0.01) for the subjects consuming the salmon diet as compared to that measured after 20 days of the stabilization diet. Mean platelet volume increased significantly during the period in which the volunteers consumed the salmon diet compared to the baseline diet (p<0.01), while the mean platelet levels decreased. Platelet aggregation (PA) was measured in platelet rich plasma before, during, and after the salmon diet using collagen, ADP, arachidonic acid (AA), and thrombin agonists. The PA threshold for ADP was significantly increased for the subjects on the salmon diet (p<0.05). No change in the PA threshold was detected for collagen or thrombin. The PA threshold for AA was unchanged also, but the platelets in subjects consuming the salmon diet had a prolonged time to maximum aggregation (p<0.01) with this reagent compared to platelets from men on the stabilization diet. Plasma, red cell, and platelet total FA composition was determined by capillary GLC. While the men consumed the salmon diets, there were marked increases (3 to 10-fold) in the EPA and DHA levels in all blood components with concomitant decreases in linoleic acid and AA levels. Thus, a salmon diet, high in n−3 FA, did not influence the bleeding times, but it decreased the sensitivity of platelets to ADP and AA, increased the mean platelet size, decreased the platelet count, and changed the FA composition of the plasma, RBC and platelet membrane lipids.

Inactivation of Viruses, Bacteria, Protozoa, and Leukocytes in Platelet Concentrates: Current Research Perspectives

LTHOUGH increased safety of blood components has been achieved through continued improvements in donor testing, concern remains about the safety of blood products. A recent government report estimated that 2.7 of every 1,000 patients who receive a transfusion of 5 units of blood components are at risk of acquiring an infectious disease. 1 Currently, prevention of infectious disease transmission associated with the transfusion of cellular components rests largely on pretransfusion donor evaluation and laboratory testing. Despite improvements in testing, the aggregate risk of acquiring a viral infectious disease from a donor passing all current serological testing remains 1 per 34,000 donbr exposures. 2 Transfusion of cellular components has been implicated in transmission of viral, bacterial, and protozoan diseases. 3 Although it is commonly recognized that hepatitis B virus (HBV), hepatitis (2 virus (HCV), cytomegalovirus (CMV), and the retroviruses, such as human immunodeficiency virus (HIV) and the human lymphotrophic viruses (HTLV), can be transmitted through cellular components, other pathogens are emerging as potentially significant transfusion-associated infectious agents. For example, transmission of protozoan infections attributable to trypanosomes 4-6 and babesia have been reported. 7,8 In addition to viral and protozoal infectious agents, bacterial contamination o f platelet concentrates continues to be reported 9,1~ and may be underreported. 11 More importantly, new infectious agents, for example, hepatitis G virus, may enter the donor population long before tests adequate to maintain consistent safety of the blood supply can be implemented. 12 Even when identified early, the clinical significance of new pathogens may not be fully recognized until large epidemiological studies are completed. 13 Over the past 10 years, investigation of methods for inactivation of infectious pathogens in cellular

Bacterial contamination of platelet components: potential solutions to prevent transfusion-related sepsis

Bacterial contamination of platelet components (PC) is the most prevalent risk for transfusion-transmitted infection. Based on the recent studies with optimal culture methods of expired PC, the prevalence of bacterial contamination is estimated to occur in approximately one in 750 to one in 1000 PC. Only within the last few years have the magnitude of the risks and the range of clinical outcomes associated with bacterial contamination been extensively characterized. Despite increased recognition of bacterial contamination of PC, transfusion-related sepsis is infrequently reported. This has largely been attributed to passive reporting systems, and low levels of clinical awareness for transfusion-related sepsis by primary care physicians. The risk for transfusion of contaminated PC has generally been characterized per component. Importantly, because patients require repeated transfusions of PC during a period of transfusion-dependent thrombocytopenia, it is appropriate to express the risk to receive a contaminated PC on a patient exposure basis. Assuming that the average hematology oncology patient may receive seven PC during a 28-day period of support, the risk of exposure to a contaminated PC is in the range of one in 150 per patient. This level of risk would not be acceptable for other intravenous medications. With increased appreciation of the risk of bacterial contamination, methods were developed to limit the risk of transfusion-transmitted bacteremia. This article focuses on those interventions that have been implemented in routine practice. The most important methods employed to mitigate the risk are improved skin disinfection, initial blood draw diversion, bacterial detection and pathogen inactivation/reduction. These technologies are now undergoing increased use in the clinical practice of transfusion medicine. With increased use, additional data are being generated to more fully characterize the effects of these interventions. Improved disinfection, blood diversion and bacterial detection have decreased, but not resolved the risk of bacterial contamination. Pathogen inactivation/reduction offers the potential for a further substantial decrease of the risk for transfusion of PC contaminated with bacteria.

INTERCEPT plasma: comparability with conventional fresh‐frozen plasma based on coagulation function – an in vitro analysis

Backgroundu2002 An effective pathogen inactivation (PI) technology for plasma must inactivate a broad range of pathogens with retention of haemostatic function suitable for therapeutic support. This study evaluated a broad panel of coagulation factors regarding functionality in plasma treated with the INTERCEPT Blood SystemTM (I‐FFP).

Therapeutic efficacy of pooled buffy‐coat platelet components prepared and stored with a platelet additive solution

Summary.u2002 Despite the introduction of platelet additive solutions for the preparation of pooled platelet components, only a few studies of limited scope have evaluated the clinical efficacy of platelets stored in these solutions. The current report presents an analysis of data to evaluate the response to the transfusion of pooled buffy‐coat components suspended in storage solution with reduced (35%) plasma content in comparison with 100% plasma products.

Structural characterization and functional effects of a circulating heparan sulfate in a patient with hepatocellular carcinoma

A circulating anticoagulant was isolated from the plasma of a 42‐year‐old man with cirrhosis and hepatocellular carcinoma who had an unusual coagulation test profile. The patient developed a fatal coagulopathy, unresponsive to protamine therapy or plasma exchange following liver biopsy. However, at presentation, routine hemostasis assays were normal. The patient had mucocutaneous bleeding but the sole laboratory abnormality was a prolonged thrombin time (TT = 99 s, normal 25–35 s). Protamine titration indicated activity equivalent to a heparin concentration of 6–7 U/ml. Antithrombin III (AT III) antigen and activity were markedly elevated. The anticoagulant activity, purified from plasma by DEAE chromatography, was identified as a glycosaminoglycan (GAG). GAG anti‐thrombin activity was completely abolished by heparin lyase III. Based on the degree of sulfation and HPLC pattern, the GAG was classified as heparan sulfate. Low levels (4 μM) of purified GAG markedly prolonged the TT (>120 s) but not the activated partial thromboplastin time (PTT) (31.4 s). In a Factor Xa assay, the GAG exhibited a potency equivalent to 0.06 U of low molecular weight heparin per nmol of uronic acid. Patients with endogenous circulating glycosaminoglycans can present with unusual laboratory coagulation test profiles. These reflect complex dysfunction of hemostasis, leading to difficulty in providing diagnosis and effective care. Am. J. Hematol. 58:285–292, 1998.

How much do we know about the platelet transfusion threshold

H ow solid is the evidence on which we base decisions regarding platelet transfusion therapy? In this issue of TRANSFUSION, Heddle and colleagues1 re-examine a critical point in transfusion medicine: how do we make decisions about what we do? In this case, at what platelet count do we prescribe prophylactic platelet transfusion? Each year approximately 220,000 patients in the United States receive 1,800,000 platelet transfusions. Prior to the advent of widely available platelet transfusion, severe hemorrhagic outcomes contributed significantly to the morbidity and mortality of thrombocytopenic patients.2 Among the blood components that we collect and prepare, platelets are the most costly and the most scarce. Thus, it is rational to examine the decisionmaking process for when to transfuse. For hematologyoncology patients the majority of platelet transfusions are prophylactic, given when the platelet count falls below a specified threshold. However, until the last decade, the issue of what threshold was appropriate had not been critically questioned.3 From 1991 to 2001, a series of studies were reported resulting in the suggestion to adjust the platelet transfusion threshold downward from 20 109 to 10 109/L.4–10 Heddle and colleagues have reviewed these studies and offer commentary regarding the design and conduct of these and future studies. They stimulate our thinking about how we interpret the data from these published studies on which we have changed our practice. Over the last 5 years these studies have had a substantial impact on downward adjustment of the threshold, but how robust were the studies that supported this change in practice? The questions posed are especially relevant and challenging because for many years clinicians have used the posttransfusion platelet count or the count increment (frequently termed the “platelet bump”) and rarely the CCI as a surrogate measure of therapeutic response. Relatively few studies have examined bleeding as the primary outcome variable to evaluate either the transfusion threshold or the dose with respect to platelet transfusion efficacy. How well defined is the relationship between the count increment and the prevention of bleeding? The observations of Heddle et al.1 regarding the conduct of the studies, their suggestions about improvements in study design, and by inference the shortcomings of the studies that provided the basis for the platelet transfusion threshold shift should provoke us to reexamine the data on which the decision to adjust the threshold was made. The first point to note is that only two prospective, randomized clinical trials have been conducted to address the issue of the appropriate platelet transfusion threshold.6,9 The first study involved 255 patients and the second 78 patients; both studies focused on patients with acute leukemia. Neither of these studies was designed as an equivalence study to test a hypothesis as to whether the two thresholds were equivalent for prevention of bleeding. The power of these studies to discern equivalence or a difference in bleeding was dependent on the estimated incidence of bleeding. Only one of the studies estimated the proportion of patients expected to develop bleeding prior to the study with reference to the potential power of the study.9 Neither study formally indicated the power to detect predefined differences between the treatment groups. Given the difference in the size of these two studies, it seems likely that they had very different power, and it is likely that the smaller study was underpowered to detect meaningful differences in bleeding. The remaining five studies were observational, ranging in size from 48 to 190 patients who were assigned randomly between two4,7,8,10 or three5 treatment groups. The latter study crossed patients over between different platelet transfusion thresholds, thus potentially complicating the analysis with respect to prophylaxis against bleeding during repeated platelet transfusions. Most likely, none of these studies was highly powered to detect clinically relevant differences or equivalence between the thresholds. In the course of their review, Heddle et al.1 emphasize several important issues relating to the design of bleeding studies: the need for defined, validated, objective bleeding assessment tools; the danger of observational bias; and the choice of analytical methods. They point out five different measures that have been used in the studies reviewed: 1) the proportion of patients with bleeding, 2) the cumulative frequency of bleeding during the period of thrombocytopenia, 3) the severity of bleeding expressed as a bleeding grade, 4) the duration each patient was observed, and 5) the temporal relationship between bleeding events. Assessment of bleeding is complex because bleeding events may not be discrete in time. Resolution of one bleeding event followed by commencement of another may not be discernible. In addition, TRANSFUSION 2003;43:691-693.