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Human leucocyte antigen (HLA) sensitisation, including the formation of antibodies against HLA, can cause serious effects in patients receiving blood. Under certain circumstances, donor HLA antibodies in the blood product can trigger the patient's granulocytes to release mediators that cause transfusion-associated lung injury (TRALI), a serious complication of transfusion. The HLA systems of both donor and patient are involved in transfusion-associated graft-versus-host disease, which is a rare disease with a high mortality. Patient HLA antibodies can destroy incompatible platelets and may cause refractoriness to platelet transfusion. Identification of a patient's HLA antibody specificities is necessary for issuing compatible platelets to overcome refractoriness. Many techniques for the detection and identification of HLA antibodies have been developed, including complement-dependent cytotoxicity assay, bead-based assays, the platelet adhesion immunofluorescence test, and the monoclonal antibody-specific immobilisation of platelet antigens assay. Different strategies for the selection of HLA-compatible platelets are applied. These strategies depend on the breadth of antibody reactivity and range from avoiding single HLA antigens in the platelet concentrates issued to apheresis of platelets from HLA-identical donors. The mechanisms of HLA sensitisation and the efforts made to provide compatible blood products to sensitised patients are reviewed in this article from the perspective of clinical transfusion medicine., Competing Interests: C. Weinstock received honoraria from Grifols, S.A. M. Schnaidt has no conflicts of interest to declare., (Copyright © 2019 by S. Karger AG, Basel.)

There is unequivocal evidence that alpha-synuclein plays a pivotal pathophysiological role in neurodegenerative diseases, and in particular in synucleinopathies. These disorders present with a variable extent of cognitive impairment and alpha-synuclein is being explored as a biomarker in CSF, blood serum and plasma. Considering key events of aging that include proteostasis, alpha-synuclein may not only be useful as a marker for differential diagnosis but also for aging per se. To explore this hypothesis, we developed a highly specific ELISA to measure alpha-synuclein. In healthy males plasma alpha-synuclein levels correlated strongly with age, revealing much lower concentrations in older (avg. 58.1 years) compared to younger (avg. 27.6 years) individuals. This difference between the age groups was enhanced after acidification of the plasmas (p<0.0001), possibly reflecting a decrease of alpha-synuclein-antibody complexes or chaperone activity in older individuals. Our results support the concept that alpha-synuclein homeostasis may be impaired early on, possibly due to disturbance of the proteostasis network, a key component of healthy aging. Thus, alpha-synuclein may be a novel biomarker of aging, a factor that should be considered when analyzing its presence in biological specimens.

Background: Substantial progress in human leukocyte antigen antibody specification has been made by the introduction of Luminex single-antigen bead (SAB) assays. This progress was impaired when it turned out that this method is prone to a prozone effect leading to false-negative results in the case of high antibody titers. Testing serum and ethylenediaminetetraacetic acid (EDTA) plasma of one patient in parallel, we observed the prozone effect with the serum sample only. This led us to investigate complement component 1 (C1) as the cause of the prozone in SAB testing. We also found an easy way to overcome the prozone effect., Methods: Sera with a prozone effect were tested in the SAB assay, applying different methods of serum pretreatment to explore the parameters leading to the prozone., Results: The prozone was not observed when EDTA plasma or serum with EDTA added were tested. Further, addition of dithiothreitol, addition of C1 inhibitor, or heat inactivation of the sera abolished the prozone effect. Adding fresh nonimmune serum to heat-inactivated sera restored the prozone effect. Only beads showing a prozone were found to be covered with C1q., Conclusion: Our observations are consistent with the hypothesis that dissociation or destruction of complement C1 eliminates the prozone effect. Addition of EDTA to serum of highly immunized patients is the easiest way to avoid false-negative results in SAB testing caused by a prozone effect.

Incompatible blood group antigens are highly immunogenic and can cause graft rejections. Focusing on distinct carbohydrate- and protein-based membrane structures, defined by blood group antigens, we investigated human bone marrow-derived mesenchymal stem cells (MSCs) cultured in human serum. The presence of H (CD173), ABO, RhD, RhCE, RhAG, Kell, urea transporter type B (SLC14A1, previously known as JK), and Duffy antigen receptor of chemokines (DARC) was evaluated at the levels of genome, transcriptome and antigen. Fucosyltransferase-1 (FUT1), RHCE, KEL, SLC14A1 (JK) and DARC mRNA were transcribed in MSCs. FUT1 mRNA transcription was lost during differentiation. The mRNA transcription of SLC14A1 (JK) decreased during chondrogenic differentiation, while that of DARC increased during adipogenic differentiation. All MSCs synthesized SLC14A1 (JK) but no DARC protein. However, none of the protein antigens tested occurred on the surface, indicating a lack of associated protein function in the membrane. As A and B antigens are neither expressed nor adsorbed, concerns of ABO compatibility with human serum supplements during culture are alleviated. The H antigen expression by GD2dim+ MSCs identified two distinct MSC subpopulations and enabled their isolation. We hypothesize that GD2(dim+) H(+) MSCs retain a better 'stemness'. Because immunogenic blood group antigens are lacking, they cannot affect MSC engraftment in vivo, which is promising for clinical applications., (Published 2011. This article is a US Government work and is in the public domain in the USA.)

Little is known about the neural mechanisms underlying top-down control of repetition priming. Here, we use functional brain imaging to investigate these mechanisms. Study and repetition tasks used a natural/man-made forced choice task. In the study phase subjects were required to respond to either pictures or words that were presented superimposed on each other. In the repetition phase only words were presented that were new, previously attended or ignored, or picture names that were derived from previously attended or ignored pictures. Relative to new words we found repetition priming for previously attended words. Previously ignored words showed a reduced priming effect, and there was no significant priming for pictures repeated as picture names. Brain imaging data showed that neural priming of words in the left prefrontal cortex (LIPFC) and left fusiform gyrus (LOTC) was affected by attention, semantic compatibility of superimposed stimuli during study and cross-modal priming. Neural priming reduced for words in the LIPFC and for words and pictures in the LOTC if stimuli were previously ignored. Previously ignored words that were semantically incompatible with a superimposed picture during study induce increased neural priming compared to semantically compatible ignored words (LIPFC) and decreased neural priming of previously attended pictures (LOTC). In summary, top-down control induces dissociable effects on neural priming by attention, cross-modal priming and semantic compatibility in a way that was not evident from behavioral results.

Background: Haploidentical bone marrow transplantation with preexisting anti-HLA antibodies is associated with a high risk of graft failure., Methods: A 27-year-old female patient with chronic myeloid leukemia and evidence of several osseous chloromas had no suitable matched bone marrow donor, and fluorescence cytometric cross-match (FCXM) revealed antibodies against donor-specific HLA-molecules. Immunoadsorption onto staphylococcal protein A was applied to remove these antibodies, and peripheral stem cell transplantation was performed from her haploidentical sister after a negative FCXM was documented after immunoadsorption and conditioning treatment., Results: FCXM for donor lymphocytes and stem cells remained negative throughout the posttransplant period, and engraftment of donor cells was documented on day +69., Conclusion: Immunoadsorption onto protein A should be considered in stem cell transplantation even from an haploidentical donor where anti-HLA antibodies and a positive FCXM are documented.

The reported decrease of platelet serotonin receptors in patients with migraine could be due to an autoimmune reaction. We therefore, examined sera from 42 migraineurs without aura, 26 migraineurs with aura, and 107 headache-free blood donors for platelet-reactive antibodies using the platelet adhesion immunofluorescence test, the NIH-lymphocytotoxicity test, and the monoclonal antibody-specific immobilization of platelet antigens test. IgG antibodies against non-HLA class I platelet antigens were found in 9.5% of patients with migraine without aura, 7.6% of patients with migraine with aura, and in 7.5% of controls; IgM antibodies were found in 11.9% of patients with migraine without aura, in 30.8% of patients with migraine with aura, and in 13.1% of controls. Most antibodies were directed against glycoprotein complexes II-III (fibrinogen receptor) or Ib-IX (thrombin receptor). Two patients with migraine without aura but no patient with migraine with aura nor any control subject had IgG antibodies of unknown specificity. One patient (2.4%) with migraine without aura and two patients (7.7%) with migraine with aura, as well as 2 controls (1.9%) had IgM antibodies not further specified. The differences in frequency of platelet antibodies of antibodies of known or unknown specificity in patients with migraine without aura and migraine with aura and controls were not statistically significant. Therefore, our data do not support the hypothesis of a pathophysiologically relevant autoimmune reaction against platelet serotonin receptors in th majority of patients with migraine. We can not exclude the occurrence of antibodies against neuron-specific serotonin receptors.

Experience with the MAIPA assay for the diagnosis of platelet-reactive antibodies has shown that high-titered antibodies falsify the test results. We here demonstrate 2 cases: i) A serum with high-titered HLA antibodies (100% panel reactivity in the LCT, titer between 4,000 and 12,000), and ii) Serum with a high-titered anti-HPA-1a (titer in the MAIPA assay 1,000). In both cases, it can be demonstrated that these antibodies led to unspecific reactions. In the 1st case, they interfered with the diagnosis of additional platelet-specific antibodies. Only the use of HLA-compatible platelets allowed a correct identification. On the other hand, in the high-titered anti-HPA-1a unspecific reactions were seen with the glycoproteins Ib/IX, Ia/IIa, and beta 2-microglobulin, leading to misinterpretations. These examples demonstrate that, in the test conditions as described, a correct diagnosis of high-titered sera might only be achieved by using compatible HLA or HPA cells.

Neonatal alloimmune thrombocytopenia (NAIT) is caused by maternal immunisation against a paternal antigen on fetal platelets. The antigen involved in the majority of cases is HPA-1 a (PIA1). Usually circulating platelet alloantibodies are detectable in the mother. In this report, we present a thrombocytopenic newborn with severe hemorrhagic diathesis due to materno-fetal HPA-1 a (PIA1) incompatibility. Platelet antibodies could initially not be demonstrated in the mother's serum but became detectable after four weeks. Because of the severe and protracted course of the disease, repeated platelet substitution was necessary throughout the first two months of life.

Reactivation of recipient antibody to HLA and red blood cell antigens is described in 8 patients after allogeneic bone marrow transplantation. These IgG antibodies can be detected between day 10 and day 40 after transplantation and, in 1 patient, can be shown to be antigen-independent. We hypothesize that, induced by graft recognition of recipient antigens, antigen-independent activation of sensitized recipient B cells takes place leading to transient antibody production.

In fetal alloimmune thrombocytopenia, maternal IgG antibodies directed against platelets pass into the fetal circulation and lead to the destruction of fetal platelets. Fetal thrombocytopenia is usually first noted postnatally, but the maternal alloantibodies may lead to severe fetal haemorrhage in utero in the second or, more commonly, third trimester. Platelet-specific antibodies appear to play the major role in the pathogenesis of fetal alloimmune thrombocytopenia. Antenatal serological diagnosis is complicated by the fact that no platelet-specific alloantibodies are detectable in untreated maternal serum in about 20% of cases. Even when antibodies are detected, repeated estimation of the titre in the mother gives no indication of the severity of the fetal thrombocytopenia. Therefore, when the diagnosis of fetal alloimmune thrombocytopenia is suspected, it should be confirmed and subsequently monitored by cordocentesis. The intrauterine transfusion of compatible platelets and the administration of high dose IgG infusions to the mother and/or fetus are currently being used as approaches to treatment. Despite advances in the antenatal diagnosis and therapy of fetal alloimmune thrombocytopenia, the clinical effects on the fetus remain unpredictable.

Reactivation of platelet-reactive antibodies of donor and recipient origin is described in a patient following allogeneic BMT (donor: anti-HPA-5b; recipient: anti-HLA, anti-HPA-1b). The antibodies were detected around day 15 after BMT, peaked around day 25, and then decreased. These antibodies are interpreted as an antigen-independent reactivation of secondary B-cell responses, activated in the context of recognition of host antigens by the graft.

Alloimmunization to the platelet-specific antigen systems HPA-1, -3 and -5 was studied in 59 multitransfused patients (31 females, 16 males, 12 children) with haematologic-oncologic disorders. All patients tested had broad-reacting or multispecific HLA antibodies. Of these, 10 (17%) were found to have additional platelet-specific alloantibodies. With respect to HLA sensitization, 9 out of 10 platelet-specific alloantibodies were found in sera which showed HLA antibodies reacting with more than 95% of unselected panel cells. Determination of antibody specificity revealed 6 patients with anti-HPA-1b, one patient with anti-HPA-3a, and 2 patients with anti-HPA-1b combined with anti-HPA-3a. One patient had anti-HPA-5b; in retrospect, only the HPA-5b antibody was demonstrable before the beginning of transfusion therapy and before HLA sensitization.

In this paper we present a patient with an initially questionable history of neonatal alloimmune thrombocytopenia (NAT) due to materno-fetal HPA-1a (PLA1) incompatibility. No circulating antibodies were detectable in untreated maternal serum, but an adsorption/elution technique enabled the demonstration of the platelet-specific anti-HPA-1a (anti-PLA1) in maternal serum. Cordocentesis at 35 weeks of gestation revealed a fetal platelet count of 18 x 10(9)/l. Intrauterine platelet transfusion with HPA-1a (PLA1)-negative donor platelets was performed prior to cesarean section.

Sera of hematologic-oncologic patients were tested regularly after platelet transfusions in three test systems: lymphocytotoxicity test, platelet adhesion immunofluorescence test, and--only selected sera--in the monoclonal antibody-specific immobilization of platelet antigen test. Of 388 patients 53 (14%) had HLA antibodies 5 of these in combination with platelet-specific alloantibodies. Lymphocyte-restricted (non-HLA) reactions were observed in 20 patients, the majority of which was attributed to lymphocyte-specific auto- or alloantibodies. Sera of 27 patients showed platelet-specific reactions, usually cold-reacting autoantibodies which have no effect in vivo.

The presence of membrane-bound and circulating platelet-associated IgG (PAIgG) in nine pregnant women with hemolysis, elevated liver enzymes, and low platelet count (HELLP syndrome) was investigated. The reference group consisted of 21 patients with chronic idiopathic thrombocytopenic purpura (ITP). In the majority of patients with ITP, thrombocytopenia results from the binding of PAIgG to the platelet membrane with subsequent platelet destruction. In none of the patients with HELLP syndrome membrane-bound or circulating PAIgG was found. By contrast, membrane-bound and circulating PAIgG was detected in 15 of the 21 cases of chronic ITP (71.4%). These findings suggest that platelet destruction in patients with HELLP syndrome does not result from the binding of PAIgG to the platelet membrane. Therefore, the treatment of thrombocytopenia in this syndrome with immunosuppressants (e.g., corticosteroids), as in chronic ITP, does not seem appropriate.

Alloimmune thrombocytopenia in newborns may lead to severe bleeding. Most often this thrombocytopenia is caused by incompatibility in the PlA system (mother PlA1 negative, child PlA1 positive). If the mother produces antibodies against PlA1 which cross the placenta, they will then destroy the child's platelets. Recent investigations have shown a strong correlation between HLA-DR3 and production of antibody. Since no screening for such antibodies is performed during pregnancy the resulting thrombocytopenia in the newborn will always be a sudden event. The best therapy is transfusion of PlA1 negative platelets; if no PlA1 negative donor can be found (2.25% of all donors) platelet concentrates of the mother may be used. After diagnosis of PlA related thrombocytopenia the relatives of the mother should be tested for PlA for the following reasons: 1. to determine PlA1 negative relatives to be used as possible donors in future pregnancies, and 2. to detect PlA1 negative women of child bearing age. If pregnant these women should be HLA-DR typed and followed for anti PlA1 antibodies to estimate the risk for the child and to determine further procedures, for instance sectio caesarea to avoid intracranial bleeding during birth.

Four of five patients with idiopathic thrombocytopenic purpura (ITP) responded to high-dose immunoglobulin treatment with transitory rise in platelet count to normal values, while the fifth had only minimal improvement. In the individual case there was good correlation between serum-IgG concentration and platelet count, but an influence on platelet-associated IgG could not be demonstrated with certainty. Treatment with high-dose IgG is apparently particularly suitable for patients with ITP refractory to steroids or splenectomy, in whom bleeding complications or operative procedures make a transitory rise in platelet count desirable.