A survey to assess transfusion medicine education needs for clinicians.

OBJECTIVE: To evaluate the baseline knowledge of attending clinicians, residents and medical students in transfusion medicine (TM) at our institution in order to develop specialty-specific lectures. BACKGROUND: Physicians often receive minimal training in TM. Transfusion practices vary widely among individual physicians and across institutions. This variation leads to increased and unnecessary risk to patients and elevated costs for hospitals--problems that may be addressed through TM education. METHODS: An electronic self-administered survey was developed and administered to attending physicians, house staff and third and fourth year medical students at four hospitals. The survey consisted of 3 sections including background demographics, 14 knowledge assessment (KA) questions and opinions on educational needs. RESULTS: Two hundred and twenty-five surveys were received from physicians and medical students, of which 189 were completed and analysed. The overall mean score on the KA section was 31 · 4 ± 18 · 3% (4 · 4 ± 2 · 6 of 14 questions) correct. Significantly stronger performance was noted in the following categories: participants in the specialties pathology or hematology (adult and pediatric), all fellows and participants who report greater than 5 h of formal TM education in the past 5 years. The majority of participants believed that additional training in TM was needed for themselves as well as other physicians at all training levels. CONCLUSIONS: Additional training in TM would be beneficial to and welcomed by physicians in all training levels in all specialties. Innovative specialty-specific educational programmes need to be developed and tested to ensure efficacy and durability.

The significance of a positive DAT in thalassemia patients.

The DAT is performed for the detection of antibody or complement on the surface of RBCs. Our institution previously performed DATs on all chronically transfused thalassemia patients before each transfusion episode to detect early alloimmunization. The medical records of all thalassemia patients treated at our institution from 2004 to 2007 were reviewed to determine the significance of the high rate of positive DATs (52.5% of 80 patients). The majority of IgG-reactive DATs were associated with a nonreactive eluate (65.4% of 286 eluates performed). A positive DAT was significantly associated with splenectomy (χ² = 15.4; p < 0.001), elevated IgG levels (χ² = 26.8; p < 0.001), HCV (χ² = 20.7; p < 0.001), and warm autoantibody (χ² = 5.87; p = 0.03). Multivariate analysis revealed that only HCV (OR, 5.0; p = 0.037) and elevated IgG levels (OR, 9.0; p = 0.001) were independently associated with a positive DAT. Alloimmunized thalassemic patients were more likely to have a positive DAT than nonalloimmunized patients, but this association was not significant (OR, 2.2; p = 0.11). A positive DAT did not correlate with decreased response to transfusion, RBC survival, hemolysis, or increased transfusion requirements. Only two cases of early alloimmunization were detected by DAT among 288 DAT-positive samples studied during 4 years. This study demonstrated that the routine performance of DATs on pretransfusion specimens in thalassemic patients has limited clinical utility, and the elimination of this test will improve turnaround time and decrease costs.

CCR3- and CXCR4-mediated interactions regulate migration of CD34+ human bone marrow progenitors to ischemic myocardium and subsequent tissue repair.

OBJECTIVE: Hematopoietic progenitor cells are able to induce neovascularization of ischemic myocardium, inhibit apoptosis, and prevent heart failure. They express functional CC chemokine-binding receptor 3 (CCR3) and CXC chemokine-binding receptor 4 (CXCR4); however, the role of those receptors in migration of progenitor cells into the ischemic myocardium is unknown. METHODS: Myocardial infarction was surgically induced in athymic nude rats, and human bone marrow-derived CD34+ cells or saline was injected into the tail vein. Cell chemotaxis was studied in vitro using chemotaxis chambers with or without concomitant stimulation with eotaxin or stromal cell-derived factor-1. Cell migration into ischemic myocardium was evaluated by immunohistochemistry. CCR3 and CXCR4 antibodies or local injections of stromal cell-derived factor-1 were used to investigate the role of chemokine expression in the migration capacity of the injected cells. Morphologic analysis included evaluation of apoptosis and capillary density in the ischemic myocardium. RESULTS: Ischemic rat myocardium demonstrated induced messenger RNA expression for the CCR3-binding chemokines eotaxin, RANTES (regulated on activation, normal T expressed and secreted), and monocyte chemotactic protein-3, but not the CXCR4-binding chemokine stromal cell-derived factor-1. Migration of human angioblasts to ischemic rat myocardium was inhibited by a blocking anti-CCR3 monoclonal antibody, but not by a blocking anti-CXCR4 monoclonal antibody, which instead inhibited migration to bone marrow. Finally, intramyocardial injection of stromal cell-derived factor-1 redirected migration of human angioblasts to ischemic rat hearts, resulting in augmented neovascularization, enhanced cardiomyocyte survival, and functional cardiac recovery. CONCLUSIONS: CCR3-dependent chemokine interactions regulate endogenous migration of CD34+ progenitors from bone marrow to ischemic but not to normal myocardium. Manipulating CXCR4-dependent interactions could enhance the efficacy of cell therapy after myocardial infarction.

Pilot study of tandem high-dose chemotherapy and autologous stem cell transplantation with a novel combination of regimens in patients with poor risk lymphoma.

In an effort to improve the outcome of poor-risk lymphoma patients, we evaluated a novel regimen of tandem high-dose chemotherapy (THDC) with autologous stem cell transplantation. A total of 41 patients (median age 40 years, range 15-68 years) with poor-risk non-Hodgkin's lymphoma and Hodgkin's disease were enrolled. THDC consisted of melphalan (180 mg/m2) and escalating dose mitoxantrone (30-50 mg/m2) (MMt) for the first conditioning regimen, and thiotepa (500 mg/m2), carboplatin (800 mg/m2), and escalating dose etoposide phosphate (400-850 mg/m2), (ETCb) as the second regimen. In all, 31 patients (76%) completed both transplants, with a median time between transplants of 55 days (range 26-120). The maximum tolerated dose was determined as 40 mg/m2 for mitoxantrone and 550 mg/m2 for etoposide phosphate. The overall toxic death rate was 12%. Following high-dose chemotherapy, 10 of 24 evaluable patients (42%) were in CR. The two-year overall survival and event-free survival is 67% (95% CI, 52-81%) and 45%, (95% CI, 29-61%) for the 41 patients enrolled; and 69% (95% CI, 525-586%) and 48% (95% CI, 30-67%) for the 31 patients completing both transplants. This THDC regimen is feasible but with notable toxicity in heavily pretreated patients; its role in the current treatment of high-risk lymphoma remains to be determined.

Outcomes of unrelated cord blood transplantation in pediatric recipients.

We report results of unrelated cord blood transplants (UCBT) in 29 pediatric recipients in one center and the risk factors associated with survival. Median age: 9 years (0.5-20); diagnosis: ALL (9), AML (4), CML (1), HD (3), HLH (1), NHL (3), NBL (2); B-thal (1), FA (1), FEL (1), Krabbe (1), WAS (1), SAA (1); median follow-up: 11 months; conditioning: total body irradiation (TBI)-ablative (14), chemotherapy-ablative (6) and reduced intensity chemotherapy (9); GVHD prophylaxis: MMF/FK506 (18), cyclosporin A (CsA)+steroids+/-MMF (7) or CsA+methotrexate (MTX) (4); median total nucleated cells (TNC): 3.8 x 10(7)/kg (1.1-11); median CD34+: 2.3 x 10(5)/kg (0.2-9.9); and HLA match: 2 (6/6), 5 (5/6), 22 (4/6). Neutrophil engraftment by cumulative incidence curves 63% (median 28 (95% confidence interval (CI) 18-32)). Probability of >/=grade II acute graft-versus-host disease (aGVHD) by day +60 27%, >/=grade III aGVHD 20% and chronic graft-versus-host disease 3%. Estimated 1-year overall survival (OS) 46% (95% CI 30-71) and standard risk 60% (95% CI 29-100%). Variables associated with improved survival by multivariate analysis include non-TBI-ablative conditioning (P=0.024), CD34+/kg (P=0.038) and gender (P=0.048). These results suggest that CD34/kg cell dose and non-TBI-ablative conditioning may be important variables influencing OS following UCBT in pediatric recipients. Given the small number of patients, these results should be viewed cautiously.

A pilot study of reduced intensity conditioning and allogeneic stem cell transplantation from unrelated cord blood and matched family donors in children and adolescent recipients.

Reduced intensity (RI) allogeneic stem cell transplantation (AlloSCT) was initially demonstrated in adults following HLA-matched family and unrelated adult donor AlloSCT. There is little information about RI AlloSCT in children. We report results of a pilot study of RI AlloSCT in 21 recipients (< or =21 years). Age: median 13 (0.5-21) years, 8F:13M, 14 unrelated cord blood units (UCB) (10 4/6, 4 5/6), two related BM (6/6, 5/6), four related PBSC (2 6/6, 2 5/6), and one related BM+PBSC (6/6). RI: fludarabine, busulfan (n=14); fludarabine, cyclophosphamide (n=4); fludarabine, melphalan (n=1); total body irradiation, fludarabine, cyclophosphamide (n=1); or fludarabine, cyclophosphamide, and etoposide (n=1). Graft-versus-host disease prophylaxis: FK506 0.03 mg/kg/day and mycophenolate mofetil 15 mg/kg/q 12 h. UCB median nuc/kg and CD34/kg was 4.3 x 10(7)/kg (0.9-10.8) and 1.9 x 10(5)/kg (0.3-6.9), and related BM/PBSC median nuc/kg and CD34/kg was 8.3 x 10(8) (4.7-18.9) and 5.0 x 10(6)/kg (4.6-6.4). Maximal chimerism following unrelated cord blood transplantation, 100% x 7, 98% x 1, 95% x 2, 55% x 1, and 0% x 3; related PBSC/BM, 100% x 5, 65% x 1, and 55% x 1. Graft failure occurred in 5/21 (24%). In summary, RI AlloSCT in children is feasible and tolerable (< or =25% GF) and results in > or =85% of recipients initially achieving > or =50% donor chimerism.

Arousal dissociates amygdala and hippocampal fear responses: evidence from simultaneous fMRI and skin conductance recording.

The experience and appraisal of threat is essential to human and animal survival. Lesion evidence suggests that the subjective experience of fear relies upon amygdala-medial frontal activity (as well as autonomic arousal), whereas the factual context of threat stimuli depends upon hippocampal-lateral frontal activity. This amygdala-hippocampus dissociation has not previously been demonstrated in vivo. To explore this differentiation, we employed functional magnetic resonance imaging (fMRI) and simultaneous skin conductance response (SCR) measures of phasic arousal, while subjects viewed fearful versus neutral faces. fMRI activity was subaveraged according to whether or not the subject evoked an arousal SCR to each discrete face stimulus. The fMRI-with arousal and fMRI-without arousal data provided a distinct differentiation of amygdala and hippocampal networks. Amygdala-medial frontal activity was observed only with SCRs, whereas hippocampus-lateral frontal activity occurred only in the absence of SCRs. The findings provide direct evidence for a dissociation between human amygdala and hippocampus networks in the visceral experience versus declarative fact processing of fear.

Transcript profiling of human dendritic cells maturation-induced under defined culture conditions: comparison of the effects of tumour necrosis factor alpha, soluble CD40 ligand trimer and interferon gamma.

Using cDNA arrays, we characterized patterns of gene expression in populations of human dendritic cells (DCs) produced for clinical use. Culture and maturation induction of myeloid adherent cells under serum-free conditions yielded DCs with phenotypes similar to those described in serum-based systems. Analysis of gene expression in DCs treated with tumour necrosis factor alpha, soluble CD40L trimer or interferon gamma, however, showed specific patterns for each factor examined. Our studies document the expression of several transcripts that have not hitherto been described in DCs and/or differentially regulated according to the differentiation state of the DCs, and suggest important functional differences among the DC populations examined. In addition, DC maturation directs changes in the levels of mRNA specific for transcriptional regulators that effect the production of cytokines (e.g. BCL-6, c-rel). Other changes observed, including alteration in the gene expression profile of adhesion molecules and chemokine receptors such as CD44H, CD 49B, Rantes R, CXCR5 and CD37, suggest differences in trafficking potential between the populations studied. This broad-based description of DC populations, produced under serum-free conditions, has enabled us to better define intermediate stages of DC maturation as well as the differentiation-inducing effects of cytokines on these cells.

Human leukocyte antigens (HLA)-Cw as prognostic indicators in autologous transplantation for lymphoma.

The human leukocyte antigens (HLA) function as transplantation antigens and as markers in disease association. Disparity at the HLA A, B, Cw, and DR loci in allogeneic stem cell transplants results in an increased incidence of graft-versus-host disease, graft rejection, and decreased survival. HLA class I loci A, B, and Cw also function as ligands for natural killer (NK) cell receptors in an interaction that predominantly inhibits cytolysis of target antigens. This HLA-NK cell inhibitory function is required for protection against auto-aggression, and is of unclear significance in other clinical settings. Furthermore, the prevention of auto-aggression is HLA molecule specific as demonstrated by the association of specific HLA types with autoimmune diseases. It is not known whether the HLA molecules might serve as markers for outcome in autologous transplants. We investigated an association of HLA class I molecules and early transplant outcome in a cohort of patients who underwent autologous transplantation for the treatment of lymphoma. In this retrospective study, HLA class I molecules were analyzed to determine whether they affect transplant outcome. HLA typing was performed by microlymphocytotoxicity assays. Factors such as age, sex, disease type, lactate dehydrogenase (LDH), cell dose, type of graft, and transfusion events were reviewed. Outcome was defined as death (or survival) at 6 months from the date of transplant. HLA-Cw8 was significantly associated with poor outcome (odds ratio = 18 and 9.3, p = 0.01 and 0.02 in homozygous and all patients, respectively). The HLA-A and B locus molecules were not associated with outcome. Age, sex, elevated LDH, and cell dose were not associated with outcome. A blood progenitor cell dose of greater than 6 x 10(8) nucleated cells/kg was favorably associated with outcome (p = 0.08). The number of transfusions received was not associated with outcome. In the multivariate analysis of HLAs and factors associated with outcome, HLA-Cw8 emerged as an independent risk factor for poor outcome (p = 0.03) following autologous transplantation in lymphoma patients. The association of HLA-Cw molecules with outcome in this study group indicates a need for further investigation of the HLA-mediated interactions that affect antitumor cytotoxicity, cytokine release, and regimen related toxicity.

The neural correlates of orienting: an integration of fMRI and skin conductance orienting.

In fMRI studies, the averaging of neural activity across multiple trials might obscure important psychophysiological subprocesses. The orienting response (OR) is a distinctive subprocess signalling the active orientation of attention towards potentially significant events. We sought to elucidate fMRI activity associated with visual stimuli that did or did not evoke simultaneously recorded electrodermal ORs (using customised skin conductance recording). 'With-OR' stimuli were associated with significant activity in the hippocampus, anterior cingulate and ventromedial prefrontal cortex. Averaged analysis revealed activity only in the expected visual circuits. Our results suggest that potentially significant stimuli (with-OR) activate different functional networks to familiar (without-OR) stimuli, and that orienting may therefore be an informative subprocess to consider in cognitive fMRI studies.

An immunological syndrome featuring transverse myelitis, Evans syndrome and pulmonary infiltrates after unrelated bone marrow transplant in a patient with severe aplastic anemia.

A patient with severe aplastic anemia underwent a matched unrelated bone marrow transplant, following which he developed a complex autoimmune syndrome. This featured transverse myelitis, immune mediated Coombs positive hemolytic anemia and immune thrombocytopenia (Evans syndrome), pulmonary infiltrates, eosinophilia, muscle pains and cramps and lichenoid dermatitis all of which may represent manifestations of graft-versus-host disease as they showed response to immunosuppression. Thus, although immune-mediated cytopenias after an allogeneic bone marrow transplant are rare, they should be considered as a possible cause of cytopenia in post-transplant patients.

Comparative analysis of HLA-B 8101 by serology, isoelectric focusing, and sequence-based typing.

We describe a bone marrow donor evaluation on a twenty-four-year-old patient from Honduras, Central America. The patient's phenotype included a HLA-B7v, with serologic reactivity to HLA- B7, 8101, and 48, and was consistent with, HLA-B*8101. One-dimensional isoelectric focusing (IEF) identified HLA-B 7.1, a low frequency isotype that has previously been associated with the HLA-B7 variant, B*0705. To further classify this allele, sequence based typing (SBT) was performed, confirming HLA-B*8101 sequence homology. The diagnostic evaluation of this case illustrates the correlation of sequenced based typing and isoelectric focusing in defining HLA Class I antigen characteristics and the use of IEF in screening for rare and novel alleles.

A pilot study of allogeneic bone marrow transplantation using related donors stimulated with G-CSF.

Growth factor administration to donors prior to bone marrow (BM) harvesting results in an enrichment of the graft for myeloid precursors. In animals, growth factor-primed BM has a higher repopulating ability than untreated BM. Ten patients received an HLA-identical sibling, allogeneic transplant using granulocyte colony-stimulating factor (G-CSF)-stimulated BM. Stimulation consisted of G-CSF at 10 microg/kg/day for 2 days prior to harvest. Patients were transplanted for various benign and malignant hematological conditions. The GVHD prophylaxis consisted of cyclosporine, methotrexate and/or prednisone. Compared to untreated historical control BM, stimulated BM infusions contained similar number of nucleated cells (mean +/- s.d.: 3.5 +/- 1.5 vs 4.0 +/- 0.9 x 10(8)/kg), CD34+ cells (mean +/- s.d.: 7.5 +/- 3.0 vs 9.4 +/- 6.7 x 10(6)/kg), and CD3+ cells (mean +/- s.d.: 129 +/- 30 vs 190 +/- 59 x 10(6)/kg) but higher numbers of granulocyte-macrophage colony-forming units (mean +/- s.d.: 20 +/- 12 vs 96 +/- 34 x 10(4)/kg). Patients receiving stimulated BM had prompt and stable engraftment of white cells and platelets. On average they attained an ANC of > or = 1 x 10(9)/l 9 days earlier and a platelet count of > or = 20 x 10(9)/l 6 days earlier than historical controls receiving unstimulated HLA-identical sibling BM. Hospitalization was shortened by a mean of 10 days and transfusion requirements were modest. None of the patients developed severe GVHD or disease relapse. Two patients died of severe VOD post-BMT and thus were unevaluable for platelet engraftment. A third patient died of TTP on day 76 post-BMT. Seven patients are alive and well 49-585 days post-BMT. Stimulated BM may provide a valuable alternative to allogeneic BM and PBSC transplants. Ideal stimulation regimens need to be investigated.

Plasmapheresis in primary dysfunction of hepatic transplants.

BACKGROUND: Primary dysfunction is a failure of graft function which occurs in approximately 5% of transplanted livers. Retransplantation is often required. The presence of elevated serum cytokines interleukin 6 and tumor necrosis factor with hepatic graft dysfunction, as well as the historical benefit of plasmapheresis in fulminant hepatic failure-associated coma suggest a possible role for plasmapheresis therapy in the management of primary graft dysfunction in liver transplantation. DESIGN AND METHODS: We evaluated the effectiveness of plasmapheresis in the management of primary graft dysfunction in 18 patients who underwent orthotopic liver transplantation in this institution. Patients who were diagnosed with primary dysfunction of hepatic grafts underwent a course of four daily plasma exchange procedures. The clinical outcome, patient and graft survival, was compared to that of historical controls. RESULTS: Graft survival at 10 days was 77.7% and 76.2% and patient survival at 100 days was 83.3% and 85.7% in the plasmapheresed and control groups, respectively. The patients who underwent plasmapheresis had a higher incidence of dialysis intervention, 38% versus 19%, indicating more severe graft dysfunction. In the small number of patients compared for concomitant dialysis therapy, patient survival in the plasmapheresed group was 85.7% versus 50% (control), and graft survival was 57.0% versus 50%. Serum cytokine levels of tumor necrosis factor and interleukin 6 were reduced by 66.0% and 55.2%, respectively, with a single procedure. CONCLUSION: Plasmapheresis did not significantly effect graft survival in patients with primary graft dysfunction. An increase in patient survival in severe graft dysfunction with renal failure was noted but was not significant. Removal of elevated serum cytokines TNF and IL-6 was documented.

Successful use of an enhanced immunosuppressive protocol with plasmapheresis for ABO-incompatible mismatched grafts in liver transplant recipients.

Graft and patient survival rates after transplantation of ABO-incompatible liver allografts have been poor. We used plasmapheresis and a potent immunosuppressive regimen to control hemagglutinin levels and prevent early rejection. Ten patients who had a United Network for Organ Sharing status of 4 received ABO-incompatible allografts. Quadruple immunosuppression consisted of OKT3, Cytoxan, cyclosporine, and steroid taper; prostaglandin E-1 was administrated intravenously the first week. All patients underwent perioperative plasmapheresis to maintain hemagglutinin levels < 1:16. Patient survival was 80%; graft survival was 60% at 140-505 days. The rejection rate was 90%. Three recipients (A1-->O) lost their grafts to severe rejection at 5, 12, and 30 days after transplantation. All 3 had pretransplantation hemagglutinin levels > or = 1:100. Elevated hemagglutinin levels preceded the diagnosis of severe acute cellular rejection; plasmapheresis failed to lower anti-A titers in these 3 patients. We conclude that in an urgent setting, lowering of preformed hemagglutinins via plasmapheresis in combination with quadruple induction immunosuppression allows liver transplantation across ABO barriers. In patients with high baseline levels of preformed hemagglutinins, the risk of subsequent graft loss may be increased and transplantation with an ABO-incompatible graft may serve as a lifesaving intermediate step.

Loss and reappearance of Rho(D) antigen on the red blood cells of an individual with acute myelogenous leukemia.

Complete loss of Rho(D) antigen from red blood cells (RBCs) of individuals with hematologic disorders, though not frequent, has been reported. This case reports the loss of D antigen on the RBCs of a patient with acute myelogenous leukemia and its reappearance when he was in remission. Loss of D antigen expression coincided with worsening clinical and cytogenetic disease. At the time of D antigen loss, the patient also had cytogenetic abnormalities in the bone marrow cells. When he was in remission, the chromosomal abnormalities were no longer detectable, and his RBCs regained D antigen expression.

Increased incidence of HLA antigen B35 in patients with chronic lymphocytic leukemia.

We and others have reported an increased incidence of chronic lymphocytic leukemia (CLL) among Ashkenazi (ASH) Jews of European origin. We performed HLA Class I typing on all 50 CLL patients seen by us and compared them with 3886 controls consisting of healthy blood donors from the New York Blood Center. Thirty of our CLL patients were ASH Jews, 17 of whom (57%) expressed the B35 antigen compared with 462 ASH controls (26%). Seven (39%) of the CLL Caucasian patients expressed the B35 antigen compared with 305 (14.5%) of the Caucasian controls. Combining the information from the ASH Jews and the Caucasians the difference is highly significant, (p = 0.0001). The summary odds ratio was 3.7. These results indicate an increased incidence of the antigen B35 amongst ASH and Caucasian patients with CLL.