An association between decreased cardiopulmonary complications (transfusion-related acute lung injury and transfusion-associated circulatory overload) and implementation of universal leukoreduction of blood transfusions.

BACKGROUND: Cardiopulmonary adverse events after transfusion include transfusion-related acute lung injury (TRALI) and transfusion-associated circulatory overload (TACO), which are potentially lethal and incompletely understood. STUDY DESIGN AND METHODS: To determine whether the incidence of TRALI and TACO was affected by leukoreduction we conducted a retrospective, before-and-after study of acute transfusion reactions for the 7years before and after introduction of universal leukoreduction in 2000, involving 778,559 blood components. RESULTS: Substantial decreases occurred in the rates of TRALI (-83%; from 2.8 cases per 100,000 components before to 0.48 after universal leukoreduction; p=0.01), TACO (-49%; 7.4 to 3.8 cases per 100,000; p=0.03), and febrile reactions (-35%; 11.4 to 7.4 cases per 10,000; p<0.0001). The incidence of allergic reactions remained unchanged (7.0 per 100,000 before and after universal leukoreduction). These outcomes were primarily attributable to decreased TRALI and/or TACO associated with red blood cell (RBC) and platelet (PLT) transfusions (-64%) with notably smaller decreases associated with fresh-frozen plasma or cryoprecipitate transfusions (-29%). The incidence of TRALI and/or TACO after 28,120 washed RBC and 69,325 washed transfusions was zero. CONCLUSION: These data suggest novel hypotheses for further testing in animal models, in prospective clinical trials, and via the new US hemovigilance system: 1) Is TACO or TRALI mitigated by leukoreduction? 2) Is the mechanism of TACO more complex than excessive blood volume? and 3) Does washing mitigate TRALI and TACO due to PLT and RBC transfusions?

Transient T cell depletion causes regression of melanoma metastases.

BACKGROUND: Cognate immunity against neoplastic cells depends on a balance between effector T cells and regulatory T (Treg) cells. Treg cells prevent immune attack against normal and neoplastic cells by directly suppressing the activation of effector CD4+ and CD8+ T cells. We postulated that a recombinant interleukin 2/diphtheria toxin conjugate (DAB/IL2; Denileukin Diftitox; Ontak) may serve as a useful strategy to deplete Treg cells and break tolerance against neoplastic tumors in humans. METHODS: We administered DAB/IL2 (12 microg/kg; four daily doses; 21 day cycles) to 16 patients with metastatic melanoma and measured the effects on the peripheral blood concentration of several T cell subsets and on tumor burden. RESULTS: We found that DAB/IL2 caused a transient depletion of Treg cells as well as total CD4+ and CD8+ T cells (< 21 days). T cell repopulation coincided with the de novo appearance of melanoma antigen-specific CD8+ T cells in several patients as determined by flow cytometry using tetrameric MART-1, tyrosinase and gp100 peptide/MHC conjugates. Sixteen patients received at least one cycle of DAB/IL2 and five of these patients experienced regressions of melanoma metastases as measured by CT and/or PET imaging. One patient experienced a near complete response with the regression of several hepatic and pulmonary metastases coupled to the de novo appearance of MART-1-specific CD8+ T cells. A single metastatic tumor remained in this patient and, after surgical resection, immunohistochemical analysis revealed MART1+ melanoma cells surrounded by CD8+ T cells. CONCLUSION: Taken together, these data indicate that transient depletion of T cells in cancer patients may disrupt the homeostatic control of cognate immunity and allow for the expansion of effector T cells with specificity against neoplastic cells. Several T cell depleting agents are clinically available and this study provides strong rationale for an examination of their efficacy in cancer patients. TRIAL REGISTRATION: NCT00299689 (ClinicalTrials.gov Identifier).

Developmental changes in soluble CD40 ligand.

OBJECTIVES: To determine if soluble CD40 ligand (sCD40L; formally CD154) levels vary with age and to identify age-dependent ranges in healthy pediatric and adult populations. STUDY DESIGN: sCD40L was measured in 25 neonates, 74 children (3 months-15 years of age) and 20 adults using an enzyme-linked immunosorbent assay. For age group comparisons, Mann-Whitney tests were performed. Correlation coefficients assessed relationships between plasma and serum sCD40L. RESULTS: Plasma sCD40L levels were higher in neonates than in all other age groups, (P <.001). All grouped pediatric plasma levels were significantly higher than in adults (P < .0001). There were no significant differences in plasma sCD40L between pediatric age groups. Serum levels were significantly higher in neonates than in any other age group (P < .0001). Pediatric and adult serum sCD40L levels were not significantly different. CONCLUSIONS: Plasma sCD40L levels are highest at birth and remain higher than those in adults throughout childhood. Reasons for such developmental changes remain to be investigated. Age-appropriate reference ranges should be used when sCD40L is being evaluated in pediatric disorders.

Lack of significant de novo HLA allosensitization in ventricular assist device recipients transfused with leukoreduced, ABO identical blood products.

Ventricular assist devices provide support for a failing heart and often serve as a bridge to transplantation. The use of these devices has also been associated with allosensitization to HLA antigens because of transfusion of blood products. Our program established a protocol mandating the use of leukoreduced, irradiated and ABO identical products, including platelets, in patients receiving initial implantations of VADs as a bridge to transplantation. Recipients were tested for anti-HLA antibodies before VAD implantation and monthly post-implantation by cytotoxicity and solid phase assays. We observed minimal de novo anti-HLA sensitization (<10%) in this population of 55 patients, each receiving a mean of 90 blood components, using this approach. No patient developed broad sensitization (PRA>50%). In conclusion, The use of leukoreduced, irradiated, ABO identical blood products abrogates broad allosensitization in this highly transfused population.

An association of soluble CD40 ligand (CD154) with adverse reactions to platelet transfusions.

BACKGROUND: Removal of stored supernatant abrogates most transfusion reactions to leukoreduced platelets (PLTs), suggesting that PLT-derived soluble mediators are involved. PLTs are the primary source of soluble CD40 ligand (sCD40L). Engagement of the receptor for CD40L induces synthesis of proinflammatory mediators including interleukin (IL)-6, IL-8, and monocyte chemotactic protein-1 (MCP-1). STUDY DESIGN AND METHODS: Supernatants from poststorage leukoreduced PLT concentrates were assayed for white cell- (IL-6, IL-8, MCP-1) and PLT-derived (sCD40L, RANTES) inflammatory mediators. These levels were correlated with clinical outcomes. RESULTS: Of 534 transfusions, there were 12 reported (2.2%) and 2 unreported reactions (0.4%)--10 febrile and 4 allergic. Transfusions with reactions had significantly higher levels of IL-6 (2.3-fold higher; p = 0.005), IL-8 (2.2-fold higher; p = 0.001), MCP-1 (2.6-fold higher; p = 0.002), and sCD40L (1.24-fold higher; p = 0.015), but not RANTES. (1.14-fold higher; p = 0.22). The vast majority (>93%) of patients transfused with mediator levels in the highest quintile had no reactions. When levels of all five mediators were summed, the reaction rates in the first through fifth quintiles increased from 1 to 7 percent (p = 0.027). All but one reaction occurred in patients with hematologic malignancies (13 reactions/380 transfusions; 3.4%; p = 0.04 vs. other diagnoses). CONCLUSIONS: These are the first data demonstrating that a PLT-derived mediator, sCD40L, is associated with adverse transfusion events. Existing clinical factors, for example, inflammation or leukopenia, may influence whether infused mediators cause reactions.

Soluble CD40 ligand accumulates in stored blood components, primes neutrophils through CD40, and is a potential cofactor in the development of transfusion-related acute lung injury.

Transfusion-related acute lung injury (TRALI) is a form of posttransfusion acute pulmonary insufficiency that has been linked to the infusion of biologic response modifiers (BRMs), including antileukocyte antibodies and lipids. Soluble CD40 ligand (sCD40L) is a platelet-derived proinflammatory mediator that accumulates during platelet storage. We hypothesized that human polymorpho-nuclear leukocytes (PMNs) express CD40, CD40 ligation rapidly primes PMNs, and sCD40L induces PMN-mediated cytotoxicity of human pulmonary microvascular endothelial cells (HMVECs). Levels of sCD40L were measured in blood components and in platelet concentrates (PCs) implicated in TRALI or control PCs that did not elicit a transfusion reaction. All blood components contained higher levels of sCD40L than fresh plasma, with apheresis PCs evidencing the highest concentration of sCD40L followed by PCs from whole blood, whole blood, and packed red blood cells (PRBCs). PCs implicated in TRALI reactions contained significantly higher sCD40L levels than control PCs. PMNs express functional CD40 on the plasma membrane, and recombinant sCD40L (10 ng/mL-1 mug/mL) rapidly (5 minutes) primed the PMN oxidase. Soluble CD40L promoted PMN-mediated cytotoxicity of HMVECs as the second event in a 2-event in vitro model of TRALI. We concluded that sCD40L, which accumulates during blood component storage, has the capacity to activate adherent PMNs, causing endothelial damage and possibly TRALI in predisposed patients.

Decreased sepsis related to indwelling venous access devices coincident with implementation of universal leukoreduction of blood transfusions.

BACKGROUND: Randomized trials and animal models demonstrate that leukoreduction of transfusions can reduce the risk of post-operative infections. We performed a retrospective study of sepsis related to indwelling venous access devices (line related infections) before and after the July 2000 implementation of universal leukoreduction. METHODS: Line related infection data were collected from hospital infection control records for an 18-month period before and after July 2000. Transfusion histories were obtained from transfusion service records. RESULTS: Line related infections decreased in number from 150 to 98 (-35%) in transfused patients after implementation of universal leukoreduction, whereas line related infections increased from 41 to 45 (+10%) in nontransfused patients (p = 0.04). This corresponded to a decrease from 5.3 to 3.3 infections/10,000 patient days in transfused patients (p = 0.002). The infection rate remained stable in nontransfused patients at 1.5 infections/10,000 patient days both pre- and postimplementation of universal leukoreduction. Quantitatively similar decreases (33-45%) were seen in transfused surgical, medical and pediatric patients. CONCLUSIONS: A substantial and statistically significant decrease in line related infections occurred coincident with implementation of universal leukoreduction. These improved outcomes were observed in transfused but not nontransfused patients, across all clinical services, suggesting a causal relationship with universal leukoreduction.

WBC reduction of RBC transfusions is associated with a decreased incidence of RBC alloimmunization.

BACKGROUND: Allogeneic transfusion stimulates Th2 (humoral) immunity. A hypothesis was developed that WBC reduction, by reducing the Th2 stimulus associated with transfusions, might reduce RBC alloimmunization. STUDY DESIGN AND METHODS: The first retrospective cohort study involved determining the prevalence of newly detected alloimmunization in transfused patients with acute myeloid leukemia (AML) in our hospital in the period before WBC reduction and after its introduction for this particular group of patients. The second study involved determining the incidence of newly detected RBC alloimmunization in all transfused hospital patients during three annual periods with WBC-reduction prevalences ranging from 0 to 100 percent. RESULTS: The alloimmunization prevalence rate in AML patients was 8.2 percent in those receiving non-WBC-reduced RBCs and platelets (n=195) and 2.8 percent in those receiving only WBC-reduced components (n=215) (p=0.016). In all patients, the alloimmunization incidence rate decreased from 3.47 per 1000 antibody screens in 1987 (no WBC reduction) to 2.97 per 1000 in 1999 (40% of transfusions WBC-reduced) to 2.38 per 1000 in 2001 (100% of transfusions WBC-reduced) (p=0.0298). A decrease in alloimmunization was observed in both males and females, with the decrease more clearly evident in males. CONCLUSION: These preliminary data support the hypothesis that WBC reduction may be associated with a reduced frequency of RBC alloimmunization. These findings require confirmation and further investigation.

Human bone marrow megakaryocytes and platelets express PPARgamma, and PPARgamma agonists blunt platelet release of CD40 ligand and thromboxanes.

Peroxisome proliferator-activated receptor gamma (PPARgamma) is a ligand-activated transcription factor important in lipid metabolism, diabetes, and inflammation. We evaluated whether human platelets and megakaryocytes express PPARgamma and whether PPARgamma agonists influence platelet release of bioactive mediators. Although PPARgamma is mainly considered a nuclear receptor, we show that enucleate platelets highly express PPARgamma protein as shown by Western blotting, flow cytometry, and immunocytochemistry. Meg-01 megakaryocyte cells and human bone marrow megakaryocytes also express PPARgamma. Platelet and Meg-01 PPARgamma bound the PPARgamma DNA consensus sequence, and this was enhanced by PPARgamma agonists. Platelets are essential not only for clotting, but have an emerging role in inflammation in part due to their release or production of the proinflammatory and proatherogenic mediators CD40 ligand (CD40L) and thromboxanes (TXs). Platelet incubation with a natural PPARgamma agonist, 15d-PGJ(2), or with a potent synthetic PPARgamma ligand, rosiglitazone, prevented thrombin-induced CD40L surface expression and release of CD40L and thromboxane B(2) (TXB(2)). 15d-PGJ(2) also inhibited platelet aggregation and adenosine triphosphate (ATP) release. Our results show that human platelets express PPARgamma and that PPARgamma agonists such as the thiazolidinedione class of antidiabetic drugs have a new target cell, the platelet. This may represent a novel mechanism for treatment of inflammation, thrombosis, and vascular disease in high-risk patients.