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ABO blood group antigens, which are complex carbohydrate moieties, and the first human polymorphisms identified, are critical in transfusion medicine and transplantation. Despite their discovery over a century ago, significant questions remain about the development of anti-ABO antibodies and the structural features of ABO antigens that cause hemolytic transfusion reactions. Anti-ABO antibodies develop naturally during the first few months of life, in contrast to other red blood cell (RBC) alloantibodies which form after allogeneic RBC exposure. Anti-ABO antibodies are the most common immune barrier to transfusion and transplantation, but the factors driving their formation are incompletely understood. Some studies suggest that microbes that express glycans similar in structure to the blood group antigens could play a role in anti-blood group antibody formation. While the role of these microbes in clinically relevant anti-blood group antibody formation remains to be defined, the presence of these microbes raises questions about how blood group-positive individuals protect themselves against blood group molecular mimicry. Recent studies suggest that galectins can bind and kill microbes that mimic blood group antigens, suggesting a unique host defense mechanism against microbial molecular mimicry. However, new models are needed to fully define the impact of microbes, galectins, or other factors on the development of clinically relevant naturally occurring anti-blood group antibodies., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: In this retrospective study, our aim was to find the effect of leucodepleted (LD) blood transfusions on the formation of anti-HLA-antibodies when compared to non-leucodepleted (non-LD) transfusions using Luminex-based method., Methods: In this study, Luminex single antigen bead assay (L-SAB) and HLA typing were performed on 310 patients. Test positivity rates (as MFI - Mean florescence intensity) were analyzed according to the different sensitization events and gender., Results: Of the 310 patients included in the study, 58.06% (180) patients were male and 41.93% (130) were female. The average age of the patients was 42.86 (±12.37) years. In this study, test positivity rates were significantly lower in the patients who received LD RBC units than in those who received non-LD RBC units (28.43% = 29 of 102 Vs 55.22% = 74 of 134, p < 0.05). In our study, transfusion combined with a history of pregnancy had higher number of significant HLA antibodies compared to cases where transfusion was the only sensitization event (81.81% = 18/22 Vs 39.71% = 85/214, p < 0.05). In addition, anti-HLA-antibodies-MFI were significantly (p < 0.01) higher in non-LD patients compared to LD patients., Conclusion: Patients who received LD RBC units had a significantly lower rate of transfusion-associated alloimmunization compared to those who received non-LD RBC units. Multiparous women had a high risk for transfusion-related alloimmunization compared to both nulliparous women and male patient. Furthermore, class I-anti-HLA-antibodies (HLA-B and HLA-A + B) were significantly associated with pregnancy sensitization and/or blood transfusion as a single sensitization., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier GmbH. All rights reserved.)

c-Myc protein encoded by c-Myc (cellular-myelocytomatosis viral oncogene) gene regulates the related gene expression through the Wnt/β-catenin signaling pathway, and has received extensive attention in recent years. The purpose of this study was to express Helicoverpa armigera c-Myc gene ( Ha-c-Myc ) by using prokaryotic expression system, prepare the polyclonal antibody, examine the spatio-temporal expression profile of Ha-c-Myc, and investigate the possible function of Ha-c-Myc in regulating H . armigera sterol carrier protein-2 (SCP-2) gene expression. The Ha-c-Myc gene was amplified by PCR and cloned into a prokaryotic expression plasmid pET-32a(+). The recombinant plasmid pET-32a- Ha-c-Myc was transformed into Escherichia coli BL21. IPTG was used to induce the expression of the recombinant protein. Protein was purified by Ni 2+ -NTA column and used to immunize New Zealand rabbits for preparing the polyclonal antibody. The Ha-c-Myc expression levels in different developmental stages (egg, larva, prepupa, pupa, and adult) of H . armigera and different tissues (midgut, fat body, head, and epidermis) of the prepupa were determined by real-time quantitative reverse transcription PCR (qRT-PCR). Ha-c-Myc siRNA was synthesized and transfected into H . armigera Ha cells. The relative mRNA levels of Ha-c-Myc and HaSCP-2 in Ha cells were detected by qRT-PCR. Results showed that the pET-32a- Ha-c-Myc recombinant plasmid was constructed. The soluble Ha-c-Myc protein of about 65 kDa was expressed in E . coli . The polyclonal antibody was prepared. Western blotting analysis suggested that the antibody had high specificity. Enzyme linked immunosorbent assay (ELISA) showed that the titer of the antibody was high. Ha-c-Myc gene expressed at all developmental stages, with high levels in the early and late instars of larva, and the prepupal stage. Tissue expression profiles revealed that Ha-c-Myc expressed in various tissues of prepupa, with high expression level in the midgut, but low levels in the epidermis and fat body. RNAi results showed that the knockdown of Ha-c-Myc expression significantly affected transcription of HaSCP-2 , leading to a 50% reduction in HaSCP-2 mRNA expression level. In conclusion, the Ha-c-Myc was expressed through a prokaryotic expression system, and the polyclonal anti-Ha-c-Myc antibody was obtained. Ha-c-Myc may promote the expression of HaSCP-2 and play an important role in the lipid metabolism of H . armigera . These results may facilitate further study on the potential role and function mechanism of Ha-c-Myc in H . armigera and provide experimental data for exploring new targets of green pesticides.

While red blood cell (RBC) transfusion is the most common medical intervention in hospitalized patients, as with any therapeutic, it is not without risk. Allogeneic RBC exposure can result in recipient alloimmunization, which can limit the availability of compatible RBCs for future transfusions and increase the risk of transfusion complications. Despite these challenges and the discovery of RBC alloantigens more than a century ago, relatively little has historically been known regarding the immune factors that regulate RBC alloantibody formation. Through recent epidemiological approaches, in vitro-based translational studies, and newly developed preclinical models, the processes that govern RBC alloimmunization have emerged as more complex and intriguing than previously appreciated. Although common alloimmunization mechanisms exist, distinct immune pathways can be engaged, depending on the target alloantigen involved. Despite this complexity, key themes are beginning to emerge that may provide promising approaches to not only actively prevent but also possibly alleviate the most severe complications of RBC alloimmunization.

Predicted Indirectly ReCognizable Human Leukocyte Antigen (HLA) Epitopes (PIRCHE) are known to be a significant risk factor for the development of donor HLA-specific antibodies after organ transplantation. Most previous studies on PIRCHE limited their analyses on the presentation of the HLA-DRB1 locus, although HLA-DRB3/4/5, -DQ, and -DP are also known for presenting allopeptides to CD4+ T cells. In this study, we analyzed the impact of predicted allopeptides presented by these additional loci on the incidence of HLA-specific antibodies after an immunization event. We considered pregnancy as a model system of an HLA immunization and observed child-specific HLA antibody (CSA) development of 231 mothers during pregnancy by samples being taken at delivery. Our data confirm that PIRCHE presented by HLA-DRB1 along with HLA-DRB3/4/5, -DQ, and -DP are significant predictors for the development of CSA. Although there was limited peptidome overlap observed within the mothers' presenting HLA proteins, combining multiple presenting loci in a single predictor improved the model only marginally. Prediction performance of PIRCHE further improved when normalizing scores by the respective presenters' binding promiscuity. Immunogenicity analysis of specific allopeptides could not identify significant drivers of an immune response in this small cohort, suggesting confirmatory studies., Competing Interests: MN works for PIRCHE AG, which develops and operates the PIRCHE web service. The UMC Utrecht has filed a patent application on the prediction of an alloimmune response against mismatched HLA. ES is listed as an inventor on this patent. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Niemann, Matern, Spierings, Schaub and Hönger.)

Background: Blood transfusion remains important in the treatment of patients with sickle cell disease (SCD). However, alloimmunization after blood transfusion is associated with patient morbidity and mortality. Triple-knockout (TKO) pigs (i.e., pigs in which the three known xenoantigens to which humans have anti-pig antibodies have been deleted) may be an alternative source of RBCs for these patients because many humans have no preformed antibodies to TKO pig RBCs (pRBCs)., Methods and Materials: In an in vitro study, plasma from alloimmunized (n = 12) or non-alloimmunized (n = 12) SCD patients was used to determine IgM/IgG binding to, and CDC of, TKO pRBCs. In an in vivo study, after an estimated 25% of blood volume was withdrawn from two capuchin monkeys, CFSE-labeled TKO pRBCs were transfused. Loss of TKO pRBCs was monitored by flow cytometry, and 7 weeks later, 25% of blood was withdrawn, and CFSE-labeled monkey RBCs were transfused., Results: The in vitro study demonstrated that plasma from neither alloimmunized nor non-alloimmunized SCD patients bound IgM/IgG to, or induced CDC of, TKO pRBCs. In the in vivo study, survival of TKO pRBCs in the two capuchin monkeys was of 5 and 7 days, respectively, whereas after allotransfusion, survival was >28 days., Conclusions: In conclusion, (1) in the present limited study, no antibodies were detected that cross-reacted with TKO pRBCs, and (2) TKO pigs may possibly be an alternate source of RBCs in an emergency if no human RBCs are available., (© 2021 AABB.)

The functional Fc gamma receptor (FcγR) IIIA polymorphism FCGR3A -V/F158 was earlier suggested to determine the potential of donor-specific HLA antibodies to trigger microcirculation inflammation, a key lesion of antibody-mediated renal allograft rejection. Associations with long-term transplant outcomes, however, have not been evaluated to date. To clarify the impact of FCGR3A -V/F158 polymorphism on kidney transplant survival, we genotyped a cohort of 1,940 recipient/donor pairs. Analyzing 10-year death-censored allograft survival, we found no significant differences in relation to FCGR3A -V/F158. There was also no independent survival effect in a multivariable Cox model. Similarly, functional polymorphisms in two other activating FcγR, FCGR2A -H/R131 (FcγRIIA) and FCGR3B -NA1/NA2 (FcγRIIIB), were not associated with outcome. There were also no significant survival differences among patient subgroups at increased risk of rejection-related injury, such as pre-sensitized recipients (> 0% panel reactivity; n = 438) or recipients treated for rejection within the first year after transplantation (n = 229). Our study results suggest that the earlier shown association of FcγR polymorphism with microcirculation inflammation may not be strong enough to exert a meaningful effect on graft survival., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wahrmann, Döhler, Arnold, Scherer, Mayer, Haindl, Haslacher, Böhmig and Süsal.)

Although de novo donor-specific anti-HLA antibodies (dnDSA) remain a barrier for human kidney transplantation (KTx), the role of regulatory T (Treg) cells in dnDSA formation remains unknown. To address this question, we evaluated Treg cell subsets in peripheral blood mononuclear cells in 15 healthy volunteers and 59 KTx recipients using flow cytometric analysis. The post-transplant CD25 high CD127 - CD4 + Treg cells in KTx recipients were down-regulated compared with those of healthy volunteers (P < .001). Among them, 11 KTx recipients showed dnDSA formation, which was associated with lower frequencies of CD25 high CD127 - CD4 + Treg cells (P = .040). Furthermore, of the total Treg cell population, CD45RA - CD25 high CD127 - CD4 + activated Treg (aTreg) cells were significantly dominant in patients with dnDSA (P = .038), but not CD45RA + CD25 high CD127 - CD4 + resting Treg cells (P = .961). In contrast, non-donor-specific anti-HLA antibody formation was not associated with CD45RA - aTreg cells (P = .772). Multivariate logistic regression analyses revealed that CD45RA - aTreg cells were independently associated with dnDSA formation (Odds ratio = 6.69, P = .040). These findings indicate that CD45RA - aTreg cells are strongly associated with dnDSA formation in KTx recipients and might be an important risk factor of antibody-mediated rejection before clinical diagnosis., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The outcome of organ transplantation is largely dictated by selection of a well-matched donor, which results in less chance of graft rejection. An allogeneic immune response is the main immunological barrier for successful organ transplantation. Donor and recipient human leukocyte antigen (HLA) mismatching diminishes outcomes after solid organ transplantation. The current evaluation of HLA incompatibility does not provide information on the immunogenicity of individual HLA mismatches and impact of non-HLA-related alloantigens, especially in vivo . Here we demonstrate a new method for analysis of alloimmune responsiveness between donor and recipient in vivo by introducing a humanized mouse model. Using molecular, cellular, and genomic analyses, we demonstrated that a recipient's personalized humanized mouse provided the most sensitive assessment of allogeneic responsiveness to potential donors. In our study, HLA typing provided a better recipient-donor match for one donor among two related donors. In contrast, assessment of an allogeneic response by mixed lymphocyte reaction (MLR) was indistinguishable between these donors. We determined that, in the recipient's humanized mouse model, the donor selected by HLA typing induced the strongest allogeneic response with markedly increased allograft rejection markers, including activated cytotoxic Granzyme B-expressing CD8 + T cells. Moreover, the same donor induced stronger upregulation of genes involved in the allograft rejection pathway as determined by transcriptome analysis of isolated human CD45 + cells. Thus, the humanized mouse model determined the lowest degree of recipient-donor alloimmune response, allowing for better selection of donor and minimized immunological risk of allograft rejection in organ transplantation. In addition, this approach could be used to evaluate the level of alloresponse in allogeneic cell-based therapies that include cell products derived from pluripotent embryonic stem cells or adult stem cells, both undifferentiated and differentiated, all of which will produce allogeneic immune responses., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ajith, Mulloy, Musa, Bravo-Egana, Horuzsko, Gani and Horuzsko.)

Some genetic and treatment-related factors are risk factors for inhibitor development in patients with hemophilia A (PwHA). However, the genotype distribution of the factor VIII gene ( F8 ) and genetic impact on inhibitor development in Japanese PwHA remain unknown. In 2007, the Japan Hemophilia Inhibitor Study 2 (J-HIS2) was organized to establish a nationwide registry system for hemophiliacs and to elucidate risk factors for inhibitor development, designed for prospective investigation following a retrospective study (J-HIS1) which had already finished. Patients, newly diagnosed after January 2007, were enrolled in J-HIS2 and followed up for inhibitor development and clinical environments since 2008 onward. In the present study, F8 genotypes of PwHA were investigated in the patients recruited from the J-HIS2 cohort as well as those with inhibitor from the J-HIS1 cohort. F8 variants identified in 59 PwHA with inhibitor in J-HIS1 were: 20 intron-22 inversions, 5 intron-1 inversions, 9 large deletions, 4 nonsense, 8 missense, 11 small in/del, and 2 splice-site variants. F8 variants identified in 267 (67 with inhibitor) PwHA in J-HIS2 were: 76(28) intron-22 inversions, 3(2) intron-1 inversion, 1(0) duplication, 8(5) large deletions, 21(7) nonsense, 109(7) missense, 40(11) small in/del, and 9(7) splice-site variants. Forty variants were novel. The cumulative inhibitor incidence rate in the severe group with null changes was 42.4% (95% confidence interval [CI]: 33.7-50.8), higher than that with nonnull changes (15.6% [95%CI: 6.8-27.8]), in J-HIS2. Relative risk for inhibitor development of null changes was 2.89. The spectrum of F8 genotype and genetic impact on inhibitor development in Japanese PwHA were consistent with the previous reports., Competing Interests: K.S. was an endowed assistant professor funded by CSL Behring. K.Y. teaches a course endowed by Takeda Co. Ltd. T.K. received research funding from Chugai Pharmaceutical Co., Ltd. and Bayer AG, and a lecture reward from Chugai Pharmaceutical Co., Ltd., Takeda Co. Ltd., Novo Nordisk A/S, Sanofi S.A., Bayer A/G, and Sysmex Corporation. K.N. has received grants from Takeda Co. Ltd. and Sanofi S.A., and research funding from Chugai Pharmaceutical Co., Ltd., F. Hoffmann-La Roche Ltd., Bayer A/G, Novo Nordisk A/S, KM Biologics, and also honoraria from Takeda Co. Ltd., Bayer A/G, Novo Nordisk A/S, Sanofi S.A., CSL Behring, Chugai Pharmaceutical Co., Ltd., F. Hoffmann-La Roche Ltd. outside the submitted work. M.T. has received research funding from Chugai Pharmaceutical Co., Ltd., F. Hoffmann-La Roche Ltd., Sanofi S.A., CSL Behring, and Novo Nordisk A/S and also honoraria from Chugai Pharmaceutical Co., Ltd., CSL Behring, Bayer AG, Takeda Co. Ltd., Sanofi S.A., and Novo Nordisk A/S outside the submitted work and listed as an entity's board of directors or advisory committee member for Chugai Pharmaceutical Co., Ltd., Novo Nordisk A/S, Sanofi S.A., and Bayer AG. K.F. has received grants and personal fees from Takeda Co. Ltd. (Baxalta/Shire), Bayer, Pfizer, CSL Behring, Novo Nordisk, Sanofi S.A. (Biogen/Bioverativ), KM Biologics (Kaketsuken) and Chugai Pharmaceutical Co. LTD, and grants from Japan Blood Products Organization, personal fees from SRL Inc., LSI Medience, Roche Diagnostics, Siemens, Sekisui Medical, Fujirebio Inc., Torii Pharmaceuticals, Octapharma, Sysmex, MSD, and other from CIMIC, outside the submitted work. A.Y. has received payment for a manuscript and consulting fee from Japan Blood Products Organization, and honoraria for lectures from Takeda Co., Ltd., Bayer A/G, and Chugai Pharmaceutical Co., Ltd. A.S. has no conflict of interest. M.S. received research funding from Chugai Pharmaceutical Co., Ltd., F. Hoffmann-La Roche Ltd., Sanofi S.A., Takeda Co. Ltd., CSL Behring, KM Biologics Co., Ltd., and Novo Nordisk A/S; and consulting fee from Chugai Pharmaceutical Co., Ltd.; and payment for lectures on speaker's bureau from Chugai Pharmaceutical Co., Ltd., Sanofi S.A., Bayer AG, and Sysmex corporation; and is listed as an entity's board of directors or advisory committee member for Chugai Pharmaceutical Co., Ltd., F. Hoffmann-La Roche Ltd., BioMarin Pharmaceutical Inc., Bayer AG, and Sanofi S.A.; and is an inventor of patents related to anti-FIXa/FX bispecific antibodies., (Thieme. All rights reserved.)

Eplet mismatches are associated with de novo DSA (dnDSA) and antibody mediated rejection (ABMR) among the general kidney transplant population. However, it is unclear whether the level of eplet mismatch can be used for risk stratification among patients with dnDSA. We performed a retrospective observational study of kidney transplant recipients with dnDSA (n = 44) transplanted between 10/2007 and 5/2014 to evaluate eplet mismatch as a risk factor for ABMR and allograft loss among dnDSA patients. High resolution typing was inferred from by imputation based on ethnicity and NMDP haplotypes, and the eplet mismatch was calculated using the Epvix algorithm. Biopsies (N = 151) from 95.3%(42/44) of patients were reviewed. The mean (SD) eplet mismatch was 69.8(22.8). The ABMR incidence was 71.4% (30/42) and 5 year death censored allograft survival was 67.4% during the mean (SD) follow-up of 5.3 (3.1) years. ABMR and death-censored allograft survival were not correlated with eplet mismatch among dnDSA patients. However, medication adherence and dnDSA MFI < 3000 were associated with reduced ABMR incidence. Among patients with both of these favorable characteristics, only 35.7% (15/42) developed ABMR. In conclusion, the level of eplet mismatch does not correlate with ABMR or allograft loss among high risk kidney transplant patients with dnDSA., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

In pre-sensitizing events, immunological memory is mainly created via indirect allorecognition where CD4 + T cells recognize foreign peptides in the context of self-HLA class II (pHLA) presented on antigen-presenting cells. This recognition makes it possible for naive CD4 + T-helper cells to differentiate into memory cells, resulting in the creation of further antibody memory. These responses contribute to effective secretion of donor-specific anti-HLA antibodies (DSA) after second encounters with the same peptide. Preformed donor-reactive CD4 + memory T cells may induce early immune responses after transplantation; however, the tools to evaluate them are limited. This study evaluated shared T cell epitopes (TEs) between the pre-sensitizing and donor HLA using an in silico assay, an alternative to estimate donor-reactive CD4 + memory T cells before transplantation. In 578 living donor kidney transplants without preformed DSA, 69 patients had anti-HLA antibodies before transplantation. Of them, 40 had shared TEs and were estimated to have donor-reactive CD4 + memory T cells. De novo DSA formation in the early phase was significantly higher in the shared TE-positive group than in the anti-HLA antibody- and shared TE-negative groups (p=0.001 and p=0.02, respectively). In conclusion, evaluation of shared TEs for estimating preformed donor-reactive CD4 + memory T cells may help predict the risk of early de novo DSA formation after kidney transplantation., Competing Interests: MN is an employee of PIRCHE AG. The UMC Utrecht has filed a patent application on the prediction of an alloimmune response against mismatched HLA. ES is listed as inventor on this patent. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Tomosugi, Iwasaki, Sakamoto, Niemann, Spierings, Nahara, Futamura, Okada, Hiramitsu, Takeda, Goto, Narumi, Watarai and Kobayashi.)

Background: Kidney transplantation is the best treatment option for end stage renal disease (ESRD), but graft rejection is still a big obstacle that occurs in spite of immunosuppressive therapy. B cells are considered as the major reason for renal graft rejection because of antibody production. Due to their roles in B cell function, we intended to evaluate the B cell activating factor (BAFF) and its receptors including BAFF receptor (BAFF-R), B cell maturation antigen (BCMA), and transmembrane activator and cyclophilin ligand interactor (TACI) in renal transplant patients., Method: The study included 40 kidney allograft patients with cAMR, 40 stable kidney allograft patients, and 8 healthy volunteers with normal kidney function. The percentage and absolute number of CD19 + B cells were analyzed by flow cytometry, the serum level of BAFF was analyzed by ELISA, and mRNA expressions of BAFF and BAFF receptors (BAFF-R, BCMA, and TACI) were measured using quantitative real-time PCR., Results: The percentage and the absolute number of B cells decreased significantly in stable and cAMR patients compared to healthy individuals. The serum level and gene expression of BAFF, as well as the mRNA level of BCMA, were increased significantly in both cAMR and stable patients compared to healthy volunteers. There was an overexpression of TACI mRNA in cAMR patients compared to stable patients., Conclusions: Both soluble protein and mRNA transcript of BAFF increased in transplant recipients. However, BAFF neither at the serum level nor at the mRNA transcript level cannot be a good biomarker for the prediction of cAMR. In addition, expression of TACI, compared to other receptors of BAFF, confers a potential to be used in distinguishing cAMR and stable kidney transplant patients., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this article., (Copyright © 2021 Shima Afzali et al.)

Antibody-mediated rejection is the principal cause of allotransplant graft failure. Available studies differ on the impact of de novo donor specific antibody (dnDSA) in pancreas transplants but are limited by patient sample size and sera sample collection. High-resolution HLA incompatibility scoring algorithms are able to more accurately predict dnDSA development. We hypothesized that HLA incompatibility scores as determined by the HLA-Matchmaker, HLA-EMMA, and PIRCHE-II algorithms would serve as a predictor of de novo donor specific antibody (dnDSA) development and clarify the role dnDSA as detrimental to simultaneous pancreas-kidney graft survival. Our results show that female sex and race were significantly associated with dnDSA development and dnDSA development resulted in worse kidney and pancreas graft survival. The majority of individuals who developed dnDSA (88%), developed anti-HLA-DQ antibody in some combination with anti-HLA class I or -DR. A multivariate analysis of the incompatibility scores showed that both HLA-Matchmaker and PIRCHE-II scores predicted anti-DQ dnDSA development. An optimal cutoff threshold for incompatibility matching was obtained for these scores and demonstrated statistical significance when predicting freedom from anti-DQ DSA development. In conclusion, increased scores from high-resolution HLA matching predict dnDSA development, and dnDSA is associated with antibody-mediated rejection and worse pancreas and kidney graft outcomes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

In recent years, the utility of vascular complement factor 4d (C4d) deposition as diagnostic tool for antibody mediated rejection (AMR) after lung transplantation, has become a controversial issue. We aimed to pinpoint the problematic nature of C4d as biomarker with a simple experiment. We quantified C4d in broncho-alveolar lavage (BAL) of lung transplant patients with diverse post-transplant complications in 3 different settings of clinically clear cases of: 1/ chronic lung allograft dysfunction (CLAD); 2/ acute complications acute rejection (AR), lymphocytic bronchiolitis (LB), antibody-mediated rejection (AMR) and respiratory infection (INF); 3/ patients with parallel C4d immunostaining and Anti-HLA. All groups were compared to BAL of stable patients. C4d was measured via standard ELISA. C4d was increased in CLAD, predominantly in RAS (p = 0.0026) but not in BOS (p = 0.89). C4d was increased in all acute events, AR (p = 0.0025), LB (p < 0.0001), AMR (p = 0.0034), infections (p < 0.0001). In patients with parallel C4d immunostaining and serum HLA antibodies, C4d was increased in C4d-/HLA- (p = 0.0011); C4d-/HLA+ (p = 0.013); HLA+/C4d + (p = 0.0081). A correlation of systemic C-reactive protein (CRP) with C4d was found in all patients (r = 0.49; p < 0.0001). We hypothesize that free C4d in BAL may only be representative of a general immune response in the transplanted lung., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Prognostic, diagnostic or predictive biomarkers are urgently needed for assessment of chronic graft-versus-host disease (cGvHD), a major risk for patients undergoing allogeneic hematopoietic stem cell transplantation. The main goal of this review generated within the COST Action EUROGRAFT "Integrated European Network on Chronic Graft Versus Host Disease" was to identify potential novel biomarkers for cGvHD besides the widely accepted molecular and cellular biomarkers. Thus, the focus was on cellular biomarkers, alloantibodies, glycomics, endothelial derived particles, extracellular vesicles, microbiome, epigenetic and neurologic changes in cGvHD patients. Both host-reactive antibodies in general, and particularly alloantibodies have been associated with cGvHD and require further consideration. Glycans attached to IgG modulate its activity and represent a promising predictive and/or stratification biomarker for cGVHD. Furthermore, epigenetic changes such as microRNAs and DNA methylation represent potential biomarkers for monitoring cGvHD patients and novel targets for developing new treatment approaches. Finally, the microbiome likely affects the pathophysiology of cGvHD; bacterial strains as well as microbial metabolites could display potential biomarkers for dysbiosis and risk for the development of cGvHD. In summary, although there are no validated biomarkers currently available for clinical use to better inform on the diagnosis, prognosis or prediction of outcome for cGvHD, many novel sources of potential markers have shown promise and warrant further investigation using well characterized, multi-center patient cohorts., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer BB declared a shared affiliation, with no collaboration, with one of the authors, FP, to the handling editor at the time of review., (Copyright © 2020 Crossland, Perutelli, Bogunia-Kubik, Mooney, Milutin Gašperov, Pučić-Baković, Greinix, Weber, Holler, Pulanić, Wolff, Dickinson, Inngjerdingen and Grce.)

Hypoxia-inducible factor (HIF) prolyl-hydroxylase inhibitors are currently used for the treatment of renal anemia. Since HIF affects immune cells, we evaluated the effect of such a drug, the roxadustat, on adaptive immunity. Cell proliferation was assessed in a two-way mixed lymphocyte reaction (MLR) with BrdU assay. In CD4 + T cells isolated from the two-way MLRs, western blotting was performed to detect the impact of roxadustat on HIF-1α and HIF-2α, the apoptotic marker cleaved caspase-3, and the master transcription factors of CD4 + T cells differentiation towards Th1, Th2, Th17, Treg and Tfh subsets. The signature cytokines of the above CD4 + T-cell subsets IFN-γ, IL-4, IL-17, IL-10, and IL-21 were measured in the supernatants. For assessing humoral immunity, we developed a suitable antibody-mediated complement-dependent cytotoxicity assay. Roxadustat stabilized HIF-1α and HIF-2α, suppressed cell proliferation, inhibited CD4 + T-cell differentiation into Th1 and Th17 subsets, while it favored differentiation towards Th2, Treg and Tfh. Roxadustat suppressed humoral immunity too. These immunosuppressive properties of roxadustat indicate that the recently introduced HIF prolyl-hydroxylase inhibitors in medical therapeutics may render the patients vulnerable to infections. This possibility should be further evaluated in clinical trials.

Ab cross-linking of HLA class I (HLA I) molecules on the surface of endothelial cells (EC) triggers proliferative and prosurvival intracellular signaling, which is implicated in the process of chronic allograft rejection, also known as transplant vasculopathy. Despite the importance of Ab-mediated rejection in transplantation, the mechanisms involved remain incompletely understood. In this study, we examined the regulation of yes-associated protein (YAP) localization, phosphorylation, and transcriptional activity in human ECs challenged with Abs that bind HLA I. In unstimulated ECs, YAP localized mainly in the cytoplasm. Stimulation of these cells with Ab W6/32 induced marked translocation of YAP to the nucleus. The nuclear import of YAP was associated with a rapid decrease in YAP phosphorylation at Ser 127 and Ser 397 , sites targeted by LATS1/2 and with the expression of YAP-regulated genes, including connective tissue growth factor ( CTGF ), and cysteine-rich angiogenic inducer 61 (CYR61). Transfection of small interfering RNAs targeting YAP/TAZ blocked the migration of ECs stimulated by ligation of HLA I, indicating that YAP mediates the increase in EC migration induced by HLA I ligation. Treatment of intact ECs with Src family inhibitors induced cytoplasmic localization of YAP in unstimulated ECs and, strikingly, blocked the nuclear import of YAP induced by Ab-induced HLA I activation in these cells and the increase in the expression of the YAP-regulated genes CTGF and CYR61 induced by HLA I stimulation. Our results identify the Src/YAP axis as a key player in promoting the proliferation and migration of ECs that are critical in the pathogenesis of transplant vasculopathy., (Copyright © 2020 by The American Association of Immunologists, Inc.)

Background: The balance between inflammatory and anti-inflammatory responses of the immune system has been demonstrated to determine the fate of transplanted allografts. Here we analyzed CD19 + CD24 hi CD38 hi immature transitional regulatory B (TRB) cells, as well as the gene and protein levels of interleukin (IL)-10 and transforming growth factor (TGF)-β in the three separate groups, include of stable transplanted subjects, chronic antibody-mediated rejection (cAMR) patients, and healthy individuals., Method: Peripheral blood mononuclear cells (PBMCs) from stable subjects (n = 36), cAMR patients (n = 36) and healthy controls (n = 18) were isolated. Flowcytometry was performed for CD19, CD24, and CD38 surface markers. ELISA and quantitative real-time PCR were performed for IL-10 and TGF-β cytokines., Result: The percentages of immature TRB cells were significantly decrease in cAMR patients (0.98%) versus stable recipients (2.81%) and healthy subjects (4.03%) (P = 0.001 and P < 0.001, respectively). Total lymphocytes, circulating B cells, memory and mature subsets of B cells did not show any significant difference between the groups. TGF-β mRNA was 3-fold upregulated in the cAMR group compared to stable patients (P < 0.001.), but without significant alteration at the protein level. Also, long-term survival renal transplant recipients had a higher protein but not mRNA levels of IL-10 than short-term survival renal transplant recipients., Conclusion: It seems that immature TRB cell subpopulation might be a crucial regulator of immune system response and plays an important role in determining the transplantation outcome. Furthermore, immunosuppressive IL-10 and TGF-β cytokines might act as a double sword and can exhibit either pathogenic or protective effects against allograft., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Introduction: The appearance of de novo donor-specific anti-human leukocyte antigen antibodies (dnDSAs) after kidney transplantation is independently associated with poor long-term allograft outcomes. The objective of the present study was to evaluate the predictive value of a flow cytometry crossmatching (FC-XM) assay after the appearance of dnDSAs related to antibody-mediated allograft rejection (ABMR) after kidney transplantation., Materials and Methods: A total of 89 recipients with dnDSAs after transplantation were included. The crossmatching results were compared with the dnDSA profile (the mean fluorescence intensity (MFI), the complement-binding activity, and the IgG subclass profile) and the biopsy's morphological features., Results: Of the 89 patients, 59 (66%) were positive in an FC-XM assay, 17 (19%) had complement-binding DSAs, 55 (62%) were positive for IgG1 and/or IgG3 in a solid phase assay, and 45 (51%) had morphological biopsy features linked to ABMR., Conclusion: An FC-XM assay was unable to discriminate between cases with or without ABMR on biopsy findings; it had a low positive predictive value (<70%) and a low negative positive predictive value (<42.9%), taking into account the sensitivity of our assay (limit of detection: DSAs with an MFI >3000). In this context, the height of the MFI of the dnDSAs might be enough for a high positive predictive value for ABMR and additional testing for complement binding activity can remain optional., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important pathway responsible for antibody-mediated rejection (AMR). Imlifidase (IdeS) cleaves human IgG into F(ab')2 and Fc fragments, potentially inhibiting ADCC. Here we examined the effect of IdeS on allo-antibody-mediated NK cell activation (Allo-CFC) and ADCC in vitro., Methods: For Allo-CFC, normal whole blood was incubated with third-party peripheral blood mononuclear cells (PBMCs) pretreated with anti-HLA antibody positive (HS) or negative (NC) sera to measure IFNγ+ NK cell%. For ADCC, normal PBMCs were incubated with Farage B (FB) cells with HS or NC sera to measure 7-AAD+ lysed FB cell%. To assess the effect of IdeS on these assays, serum-treated PBMCs (Allo-CFC-1) and serum used for PBMC pretreatment (Allo-CFC-2) in Allo-CFC, and serum used for ADCC were preincubated with IdeS. Sera from IdeS-treated patients were also tested for Allo-CFC (Allo-CFC-3)., Results: IFNγ+ NK cell% were significantly elevated in HS versus NC sera in Allo-CFC-1 (10 ± 3% versus 2 ± 1%, P = 0.001), Allo-CFC-2 (20 ± 10% versus 4 ± 2%, P = 0.01) and 7AAD+ FB cell% (11 ± 3% versus 4 ± 2%, P = 0.02) in ADCC. These were significantly reduced by IdeS treatment. Patient sera with significantly reduced anti-HLA antibody levels at 1 day postimlifidase lost the capacity to activate NK cells in Allo-CFC-3, but those at 1-3 months postimlifidase regained the capacity., Conclusions: IdeS inhibited NK cell activation and ADCC in vitro and in treated patients. These results and reported inhibition of complement activating anti-HLA antibodies by IdeS suggest its possible role in treatment of AMR.

NKG2D is a potent activating immunoreceptor expressed on nearly all cytotoxic lymphocytes promoting their cytotoxicity against self-cells expressing NKG2D ligands (NKG2DLs). NKG2DLs are MHC class I-like glycoproteins that usually are not expressed on "healthy" cells. Rather, their surface expression is induced by various forms of cellular stress, viral infection, or malignant transformation. Hence, cell surface NKG2DLs mark "dangerous" cells for elimination by cytotoxic lymphocytes and therefore can be considered as "kill-me" signals. In addition, NKG2DLs are up-regulated on activated leukocytes, which facilitates containment of immune responses. While the NKG2D receptor is conserved among mammals, NKG2DL genes have rapidly diversified during mammalian speciation, likely due to strong selective pressures exerted by species-specific pathogens. Consequently, NKG2DL genes are not conserved in man and mice, although their NKG2D-binding domains maintained structural homology. Human NKG2DLs comprise two members of the MIC (MICA/MICB) and six members of the ULBP family of glycoproteins (ULBP1-6) with MICA representing the best-studied human NKG2DLs by far. Many of these studies implicate a role of MICA in various malignant, infectious, or autoimmune diseases. However, conclusions from these studies often were limited in default of supporting in vivo experiments. Here, we report a MICA transgenic (MICAgen) mouse model that replicates central features of human MICA expression and function and, therefore, constitutes a novel tool to critically assess and extend conclusions from previous in vitro studies on MICA. Similarly to humans, MICA transcripts are broadly present in organs of MICAgen mice, while MICA glycoproteins are barely detectable. Upon activation, hematopoietic cells up-regulate and proteolytically shed surface MICA. Shed soluble MICA (sMICA) is also present in plasma of MICAgen mice but affects neither surface NKG2D expression of circulating NK cells nor their functional recognition of MICA-expressing tumor cells. Accordingly, MICAgen mice also show a delayed growth of MICA-expressing B16F10 tumors, not accompanied by an emergence of MICA-specific antibodies. Such immunotolerance for the xenoantigen MICA ideally suits MICAgen mice for anti-MICA-based immunotherapies. Altogether, MICAgen mice represent a valuable model to study regulation, function, disease relevance, and therapeutic targeting of MICA in vivo ., (Copyright © 2020 Kim, Born, Bléry and Steinle.)

Background: The aim of this study was to evaluate neutrophil extracellular traps (NETs) in kidney transplant recipients (KTR) and their potential involvement in acute antibody-mediated rejection (AAMR)., Methods: We studied 3 groups: KTR with AAMR (KTR-Cases, n = 14); KTR without any immunologic event (KTR-Controls, n = 14) and donors (n = 12). Spontaneous and lipopolysaccharide-induced NETosis were evaluated by immunofluorescence indirect (IFI) (NET/cells ratio). Plasmatic cH3-DNA complexes were evaluated by ELISA, (Optic Density Index - ODI). The expression of MPO and citrullinated histone 4 (cH4) was evaluated in renal biopsies., Results: We found an enhanced spontaneous NETosis in KTR regardless of whether they had rejection. The Nets/cells ratio in spontaneous NETosis was 0.203 (IQR 0.12-0.34) in Total-KTR and 0.094 (IQR 0.01-0.17) in donors, p = .011. Likewise, the ODI of cH3-DNA was 1.41 (IQR 0.94-1.72) in Total-KTR, and 0.95 (IQR 0.83-1.27) in donors, p = .019. KTR-Cases had the higher amount of NETs 1.70 (IQR 1.19-1.91). In two KTR-Cases, expression of MPO and cH4 was found in biopsies., Conclusions: KTR show enhanced NETosis. This may indicate a permanent activation of neutrophils. Although more studies are needed, the higher amount of NETs and netting neutrophils in biopsies of KTR-Cases suggest a role of NETosis in AAMR., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Neutrophil specific Fcγ receptor IIIb (CD16b) is a low-affinity IgG receptor. Its polymorphic variants are associated with human neutrophil antigens (HNA). HNA-1a and HNA-1b differ in four amino acids. Immunization can lead to the production of alloantibodies. The exact contribution of four amino acid exchanges for the formation of HNA-1a, -1b epitopes is currently unknown., Study Design and Methods: Permutation of each polymorphic amino acid from wild-type CD16b cDNA constructs was performed and expressed on HEK293 cells. All 16 receptor variants were produced and tested against 19 well-characterized HNA antisera in an antigen capture assay., Results: Analyzing the reaction pattern revealed that anti-HNA-1a antibodies can bind whenever asparagine (N) is present in position 65, regardless of the three other positions (CD16b *N**). Anti-HNA-1b antibodies can bind when serine (S) is present in position 36 (CD16b S***), when N is present in position 82 (CD16b **N*), or both (CD16b S*N*). CD16b variants with N65 and S36 and/or N82 (such as CD16b SNN*) bind both, anti-HNA-1a and anti-HNA-1b alloantibodies. If these specific amino acids are missing (as in CD16b RSD*), no antibodies will bind., Conclusion: Whereas the primary structure of HNA-1a and HNA-1b usually differs in four amino acids, epitope composition is not "antithetical". N65 alone determines the presence of HNA-1a, and S36 and/or N82 determine the presence of HNA-1b. Amino acid 106 does not participate in epitope formation. Our findings are of specific relevance when a HNA-1 phenotype is predicted from a genotype., (© 2020 The Authors. Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.)

Human leukocyte antigen (HLA) matching is one of the cornerstones of organ allocation in deceased-donor kidney transplantation. Increased numbers of HLA allele mismatches are associated with a higher risk of immunological rejection, de novo donor-specific HLA antibody development and graft failure. HLA epitopes are defined as the specific portions of HLA molecules to which antibodies and T-cell receptors bind with their paratopes. The same epitope can be present on different HLA alleles. Therefore, HLA matching at the epitope instead of allele level theoretically offers a more precise assessment of donor-recipient HLA compatibility and may more effectively prevent sensitization against foreign tissue. In this review, we describe the different options proposed to define clinically relevant HLA epitopes and critically discuss the potential role of HLA epitope matching in kidney transplantation., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Induction of longstanding immunologic tolerance is essential for survival of transplanted organs and tissues. Despite recent advances in immunosuppression protocols, allograft damage inflicted by antibody specific for donor organs continues to represent a major obstacle to graft survival. Here we report that activation of regulatory CD8 T cells (CD8 Treg) that recognize the Qa-1 class Ib major histocompatibility complex (MHC), a mouse homolog of human leukocyte antigen-E (HLA-E), inhibits antibody-mediated immune rejection of heart allografts. We analyzed this response using a mouse model that harbors a point mutation in the class Ib MHC molecule Qa-1, which disrupts Qa-1 binding to the T cell receptor (TCR)-CD8 complex and impairs the CD8 Treg response. Despite administration of cytotoxic T lymphocyte antigen 4 (CTLA-4) immunoglobulin (Ig), Qa-1 mutant mice developed robust donor-specific antibody responses and accelerated heart graft rejection. We show that these allo-antibody responses reflect diminished Qa-1-restricted CD8 Treg-mediated suppression of host follicular helper T cell-dependent antibody production. These findings underscore the critical contribution of this Qa-1/HLA-E-dependent regulatory pathway to maintenance of transplanted organs and suggest therapeutic approaches to ameliorate allograft rejection., Competing Interests: The authors declare no competing interest.

Objective: To evaluate the diagnostic performance of donor-derived plasma cell-free DNA (cfDNA) in discriminating antibody-mediated rejection (ABMR) or de novo donor-specific antibodies (DSA) without histological lesions in kidney allograft recipients., Methods: In this prospective single center observational study, we enrolled kidney allograft recipients between November, 2016 and September, 2017 at the First Affiliated Hospital of Sun Yat-sen University. Kidney allograft recipients with ABMR, de novo DSA but no histological lesions or negative DSA, and stable renal function were included. The plasma cfDNA fraction was measured using a targeted, single nucleotide polymorphism (SNP)-based assay. Pathological diagnosis was made according to the 2015 Banff Kidney Rejection Classification. The area under the ROC curve (AUC-ROC) was determined using the bootstrapping method to estimate median and 95% confidence interval (95% CI). The sensitivity, specificity and Youden index, positive predictive value (PPV), and negative predictive value (NPV) were calculated for specific cfDNA fractions., Results: Totally 37 consecutive patients received kidney allografts, including 18 recipients in the ABMR group and 19 recipients in the stable allograft group (7 DSA-positive and 12 DSA-negative). All patients in the ABMR group were DSA positive and 7 patients in the stable group were DSA positive but had no pathologically proven ABMR. The median donor-derived plasma cfDNA fraction was 2.4% (Q1 1.52% -Q3 3.70%) in the ABMR group, and was significantly higher than that of the stable group (0.65%, Q1 0.57% -Q3 0.97%; P < 0.001), but comparable with that of the DSA-positive patients in the stable allograft group ( P = 0.074). The AUC-ROC of cfDNA was 0.90 (95% CI, 0.79-0.98). When a cfDNA threshold of 1% was chosen, it had a sensitivity of 88.9% and a specificity of 73.7%. The PPV was 76.2% and the NPV was 87.5%., Conclusion: Donor-derived plasma cfDNA fraction increased in kidney allograft recipients with ABMR. Detection of donor-derived plasma cfDNA fraction may contribute to the discrimination between ABMR and stable renal allograft function and may aid early recognition of earlier stage antibody-mediated injury., (Copyright © 2020 Zhang, Zheng, Li, Fu, Li, Su, Zeng, Liu, Wang, Huang, Xu, Ye, Liu and Wang.)

Background: Pregnant women are routinely screened for red blood cell (RBC) antibodies early in pregnancy. If RBC-alloantibodies are detected, repeated laboratory testing is advised to timely identify pregnancies at high risk for severe hemolytic disease of the fetus and newborn (HDFN). We assessed for RBC alloantibodies, other than anti-D or anti-K, cut-offs for the titer and the antibody dependent cellular cytotoxicity (ADCC) test to select high-risk cases. To advise on test repeat intervals, and to avoid unnecessary testing, we evaluated the chance for exceeding the cut-offs for Rh antibodies other than anti-D, Jk, Fy, and S/s antibodies., Study Design and Methods: Diagnostic value of antibody titer and ADCC test was determined with data from a prospective index-cohort study, conducted in 2002-2004. Laboratory test outcomes were from a recent observational cohort (2015-2016)., Results: A titer cut-off of ≥16 showed a sensitivity of 100% (95% CI:73-100%) and a positive predictive value (PPV) of 17% (95% CI:14%-20%). The percentage of pregnancies reaching a titer above the cut-off of ≥16 varied from 0% for anti-Jk a /Jk b (n = 38) to 36% for anti-c (n = 97). The ADCC test showed no cut-off with a 100% sensitivity. However, in cases with a titer ≥16 and an ADCC test ≥30% a PPV of 38% was obtained to detect severe HDFN., Conclusion: A titer cut-off of ≥16 is adequate to detect all cases at risk for severe HDFN; the ADCC test may add a more accurate risk estimation. Repeated testing is recommended in pregnancies with anti-c. In pregnancies with other Rh antibodies a repeated test in the third trimester is recommended., (© 2019 AABB.)

Background: We conducted a meta-analysis to evaluate the predictive value of serum soluble B cell-activating factor (sBAFF) for antibody-mediated rejection (ABMR), which remains controversial., Methods: Systematic literature search was performed in PubMed, EMBASE, Scopus, Cochrane Library, Web of Science and three Chinese databases. Studies of any relevant design were included. Random and fixed-effects meta-analytical models were used. Study quality, publication bias, and heterogeneity were assessed. This study was registered with PROSPERO (CRD42019109198)., Results: Nine observational studies were included in the meta-analysis, including 1302 cases (median NOS quality score = 8, range 6-8). The incidence of ABMR was significantly higher in the high sBAFF group than in the low sBAFF level group (Risk ratio [RR] 2.04 [95% CI 1.52-2.74], I 2  = 26%, P < 0.01, N = 1014). The subgroup analysis showed that regardless of pre-transplant donor-specific antibody (DSA) status, the high sBAFF level group still had a significantly higher incidence of ABMR. sBAFF was not associated with the risk of TCMR. The sBAFF level was significantly higher in the anti-HLA-antibody (+) group than in anti-HLA-antibody (-) patients before or after kidney transplantation (Standardized mean difference [SMD] 0.43 [0.29-0.56], P < 0.01, I 2  = 34%, N = 1001)., Conclusion: sBAFF is a promising biomarker to further stratify the risk of ABMR post-renal transplant., Competing Interests: Declaration of Competing Interest The authors declared that there is no conflict of interest., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Chronic active antibody-mediated rejection is a major cause of allograft failure in kidney transplantation. Microvascular inflammation and transplant glomerulopathy are defining pathologic features of chronic active antibody-mediated rejection and are associated with allograft failure. However, the mechanisms of leukocyte infiltration and glomerular endothelial cell injury remain unclear. We hypothesized MHC class II ligation on glomerular endothelial cells (GEnC) would result in upregulation of adhesion molecules and production of chemoattractants. A model of endothelial cell activation in the presence of antibodies to MHC classes I and II was used to determine the expression of adhesion molecules and chemokines. Murine GEnC were activated with IFNγ, which upregulated gene expression of β2-microglobulin (MHC class I), ICAM1, VCAM1, CCL2, CCL5, and IL-6. IFNγ stimulation of GEnC increased surface expression of MHC class I, MHC class II, ICAM1, and VCAM1. Incubation with antibodies directed at MHC class I or class II did not further enhance adhesion molecule expression. Multispectral imaging flow cytometry and confocal microscopy demonstrated MHC molecules co-localized with the adhesion molecules ICAM1 and VCAM1 on the GEnC surface. GEnC secretion of chemoattractants, CCL2 and CCL5, was increased by IFNγ stimulation. CCL2 production was further enhanced by incubation with sensitized plasma. Endothelial activation induces de novo expression of MHC class II molecules and increases surface expression of MHC class I, ICAM1 and VCAM1, which are all co-localized together. Maintaining the integrity and functionality of the glomerular endothelium is necessary to ensure survival of the allograft. IFNγ stimulation of GEnC propagates an inflammatory response with production of chemokines and co-localization of MHC and adhesion molecules on the GEnC surface, contributing to endothelial cell function as antigen presenting cells and an active player in allograft injury., (Copyright © 2019 Elsevier B.V. All rights reserved.)

: The development of neutralizing antibodies is a rare complication of von Willebrand disease treatment. In major surgical procedures for severe forms of the disease, the recognition of ineffective therapy and alternative treatment protocols are lifesaving. We report the case of a 6-year-old girl with type 3 von Willebrand disease in whom inhibitors were sought due to ineffective haemostasis together with lower than expected von Willebrand factor (VWF) recoveries after a surgical procedure. Replacement therapy first with recombinant factor VIIa and then with high doses of recombinant factor VIII in continuous infusion successfully stopped the bleeding. A high level of anti-VWF antibodies was determined by the immunological method. A frameshift mutation associated with premature termination codon (c.2435delC, p.Pro812ArgfsTer31) was determined in our patient. Although the reports on association of this mutation with inhibitor risk are inconsistent, it represents an evidence-based diagnostic and management practice in recognition of high-risk VWF genotype.

Antibody-mediated rejection (ABMR) is an important cause of both short- and long-term injury to renal allografts. Transplant glomerulopathy (TG) is strongly associated with ABMR and reduced graft survival. Ultrastructural changes in early-stage ABMR include TG as a duplication of the glomerular basement membrane (GBM), which can be observed only by transmission electron microscopy (TEM). Low-vacuum scanning electron microscopy (LVSEM) is a new technique that allows comparatively inexpensive, rapid, and convenient observations with high magnification. We analyzed human renal transplants using LVSEM and evaluated the ultrastructural changes representing TG in ABMR. GBM duplication was more clearly visible in the LVSEM images than in the light microscopy (LM) images. In the ABMR group, the cg score of the Banff classification was higher in 54% (7/13) of specimens for LVSEM images than for LM images. And 4 specimens exhibited duplication of the GBM analyzed by LVSEM, but not by LM. In addition, three-dimensional ultrastructural changes, such as coarse meshwork structures of GBM, were observed in ABMR specimens. The ABMR group also exhibited ultrastructural changes in the peritubular capillary basement membranes. In conclusion, analyses of renal transplant tissues using LVSEM allows the identification of GBM duplication and ultrastructural changes of basement membranes at the electron microscopic level, and is useful for early-stage diagnosis of ABMR.

Over proliferation of glomerular mesangial cells (MCs) disturbs mesangial homeostasis and leads to renal damage in mesangioproliferative glomerulonephritis. It is documented that transcriptional factors may be involved in the proliferation of MCs. This study aims to identify the key transcriptional factor that prevents the MCs from over proliferation and to clarify its regulatory mechanism. Microarray analysis of glomeruli isolated from Sprague-Dawley rats (SD rats) with or without anti-Thy1 nephritis (anti-Thy1N) showed that the cell cycle pathway was the most enriched pathway in anti-Thy1N model, and the D-site binding protein (DBP) ranked first in the cluster of transcription factors. Compare with normal rats, DBP is markedly decreased accompanied by an over proliferation of MCs in rats with anti-Thy1N. The cell proliferative capacity was measured by 5-Ethynyl-2'-deoxyuridine (EdU) assay in primary rat MCs with DBP knockdown or overexpression, respectively. The results showed that the knockdown of DBP significantly promoted the proliferation of MCs, whereas the overexpression of DBP inhibited the MCs' proliferation, compared to that of the control cells. Further study indicated that DBP arrested G1/S-phase transition by inhibiting the expression of p21, p27 and inducing the Cyclin D1 expression in MCs. The current data suggest that DBP effectively inhibits the proliferation of MCs through G1 phase arrest, and the decrease of DBP may induce mesangial over proliferation in rats with anti-Thy1N., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

HLA laboratories use virtual crossmatching (VXM) to predict recipient and donor compatibility using HLA antibody data and donor HLA type. Increasingly, transplant centers are utilizing VXM as the final compatibility determination prior to transplant. However, the VXM interpretation is based on HLA experience of individual transplant centers. This study developed data-driven algorithms that predicted flow cytometric crossmatch (FCXM) outcomes using HLA antibody mean fluorescent intensity (MFI) data and donor HLA typing without the need for human interpretation.Two algorithms were evaluated; an MFI Optimal-Threshold model and a Least-Squares-Fitting model. The Optimal-Threshold model correctly determined between 81.5% and 85.5% of T or B-cell responses. A class I antibody MFI threshold of 4670 was optimal for predicting T-cell response while an antibody MFI threshold of 6180 was optimal for predicting B-cell responses. HLA class I antibodies had a 1.47-fold greater influence on FCXM outcomes than class II antibodies. HLA-B antibodies influenced T and B-cell responses more than HLA-A or -C (-B > -A > -C). The Least-Squares-Fitting model increased accuracy to 94.1% and 88.8% for T and B-cell responses, respectively. The algorithms described here provide enhanced FCXM prediction and novel insights into the influence of specific HLA antibodies on the crossmatch outcome., (Copyright © 2019 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

The occurrence of acute antibody-mediated rejection (ABMR) is higher in flow cytometric crossmatch (FCXM)-positive patients despite desensitization. Accumulating evidence suggests a correlation between the complement-binding ability of donor-specific antibodies (DSAs) and the risk of ABMR. Here, we investigated the correlation between complement C3d-fixing ability of preformed DSA and ABMR risk, the efficacy of a desensitization protocol for patients with C3d-fixing DSA, and the risk of ABMR in 21 DSA- and FCXM-positive patients. We retrospectively analyzed the C3d-fixing ability and mean fluorescence intensity (MFI) of preformed DSA before and after desensitization. Six patients had non-C3d-fixing DSA and 15 had C3d-fixing DSA. The presence of C3d-fixing DSA before desensitization was correlated with the incidence of acute ABMR within 1 year after transplantation (p = .04) and chronic ABMR (p = .03). Moreover, the MFI of preformed DSA differed between responder and non-responder C3d-fixing DSA after desensitization (p < .0001). The C3d-fixing ability of preformed DSA with low MFI disappeared after desensitization. These results indicate that measuring DSA C3d-fixing ability may identify patients with a high risk of ABMR, especially before desensitization. CLINICAL TRIAL NOTATION: UMIN Clinical Trials Registry (UMIN-CTR) number: UMIN000033449., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The negative impact of donor specific HLA alloantibodies in solid organ transplantation is well known and understood within the histocompatibility and immunogenetics community. However the influence of donor-specific antibodies in the outcome of haematopoietic stem cell transplantation is less well regarded. As donor choices have evolved from HLA matched siblings and extremely well matched unrelated donors to mismatched cord blood and haplo-identical-related donors, we are now identifying more patients with antibodies reactive against their donor mismatches. The clinical significance of the antibodies that can be detected has not yet been fully elucidated., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: HLA mismatching is a well known risk factor for worst outcomes in kidney transplantation., Methods: In the present study, HLA antigen and eplet mismatches were determined in 151 living donor-recipient pairs transplanted between 2007 and 2014 and rejection episodes and graft survival were evaluated., Results: We found that high HLA-II eplet mismatch load (EpMM ≥ 13, versus low EpMM ≤ 5), was an independent predictor of AMR (adjusted HR = 14.839; P = 0.011), while HLA-II AgMM was not. We also showed that HLA-II EpMM load was a significant better predictor of AMR than AgMM (c-statistic = 0.064; P = 0.023). After discriminating HLA-II into HLA-DR and HLA-DQ loci we demonstrated that high versus low eplet mismatch load for HLA-DR (T3 ≥ 6 versus T = 0-1, p = 0.013) and HLA-DQ (T3 ≥ 7 versus T = 0-1, p = 0.009) are independent predictors for AMR. HLA-II EpMM increased discrimination performance of the classical HLA-II AgMM risk model (IDI, 0.061, 95%CI: 0.005-0.195) for AMR. Compared with AgMM, HLA-II eplet model adequately reclassified 13 of 17 patients (76.5%) with AMR and 92 of 134 patients (68.7%) without AMR (cfNRI, 0.785, 95%CI: 0.300-1.426)., Conclusions: Our study evidences that eplet-based matching is a refinement of the classical HLA antigen mismatch analysis in LDKT and is a potential biomarker for personalized assessment of alloimmune risk., (Copyright © 2019 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Background: Platelet antibodies can lead to clinical diseases such as platelet transfusion refractoriness (PTR), fetal/neonatal alloimmune thrombocytopenia (FNAIT), etc. This study is aimed at understanding CD36 expression, platelet alloantibody distribution in different populations in Northern China, and effects of platelet alloantibodies on pregnancy., Study Design and Methods: Whole blood samples of 612 subjects including hematological patients, pregnant women, and blood donors were collected at a single center, then CD36 expressions were determined, followed by platelet antibody screening and characterization of platelet antibody specificity. A retrospective analysis was performed in 1552 pregnant women admitted to Department of Obstetrics, in order to investigate FNAIT occurrence., Results: Rate of CD36 deficiency expression was 2.12% (13/612), all cases exhibited type II deficiency without type I deficiency being detected, and such rate is lower than that in Southern China (3.43%), Japanese (4.87%) and in the black people (4.18%), and higher than that in the White people (0.09%). Positive rates of platelet antibody screening in hematological patient group (6.86%, 14/204) and in pregnant women group (6.31%, 13/206) are higher than that in blood donor group (0.49%, 1/202), P < .01. Out of 1552 pregnant women, there were not children with FNAIT., Conclusion: The frequency of CD36 deficiency in northern China was low, all of them were type II deficiency, and no CD36 antibody was detected. It is speculated that the risk of immune-related thrombocytopenia caused by CD36 deficiency in this population is very low. Platelet antibodies should be monitored early in patients with hematological and multiple miscarriages pregnant., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Aims: Chronic-active antibody mediated rejection (c-aABMR) is a major cause of kidney graft loss. Currently, little is known about the relation between histopathologic parameters and renal allograft survival., Methods and Results: Between 2008 and 2014, 41 patients with a progressive decrease in renal function were diagnosed with c-aABMR according to Banff 2015 and followed up for at least 3 years. Clinical and renal biopsy characteristics were analyzed for association with graft survival. During follow-up 26 cases lost their graft because of c-aABMR at a median follow up of 40 months after diagnosis. Cases with v-lesions in their biopsy had a significant higher loss of eGFR prior to diagnosis. The total inflammation score (r = -0.45 p = .007) and the severity of interstitial fibrosis (r = -0.38 p = .023) were related to the eGFR at time of biopsy. Univariate regression analysis showed that eGFR at time of biopsy, total inflammation, interstitial fibrosis and the sum chronicity score were significantly related to the risk for graft failure during follow-up. In a multivariate analysis only the severity of interstitial fibrosis remained associated with decreased graft survival (HR 1.9 per score point, 95% CI 1.2-2.8, p = .004)., Conclusion: Severity of renal interstitial fibrosis and not inflammation predicts graft survival in cases of c-aABMR., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Donor-specific anti-human leukocyte antigen (HLA) antibodies (DSAs) were associated with graft failure (GF) following haploidentical stem cell transplantation (Haplo-HSCT). The prevalence and risk factors of DSAs in pediatric candidates remain to be determined. In a prospective trial (ChiCTR-OPC-15006672), 486 children with hematological diseases were enrolled to screen for the presence of anti-HLA class I and II antibodies of immunoglobulin G type. Fifty two patients (10.7%) demonstrated positive panel-reactive antibody (PRA) for class I; 24 (4.9%), for class II; and 13 (2.7%), for both. Multivariate analysis showed diagnosis was the independent risk factor for antibodies, as acute lymphoblastic leukemia (ALL) patients (HR0.141, 95% CI: 0.037-0.538, p = 0.004) had a lower incidence of class II PRAs and DSAs against HLA-B, DQ, and DR, whereas myelodysplastic syndrome (MDS) patients had a higher incidence of PRAs for both class I and class II (HR4.790, 95% CI: 1.010-22.716, p = 0.049), and DSAs against HLA-A, B, C, DP, and DQ. Older age (>12 vs. ≤12) was associated with DSAs against HLA-DP (HR0.194, 95% CI: 0.041-0.918, p = 0.039). Our findings provided novel evidence for prevalence and risk factors for PRAs and DSAs in pediatric candidates receiving haplo-HSCT, possibly benefiting anti-HLA antibody monitoring and donor selection.

Fc gamma receptors (FcγRs) play a major role in the regulation of humoral immune responses. Single-nucleotide polymorphisms (SNPs) of FCGR2A and FCGR3A can impact the expression level, IgG affinity and function of the CD32 and CD16 FcγRs in response to their engagement by the Fc fragment of IgG. The CD16 isoform encoded by FCGR3A [158V/V] controls the intensity of antibody-dependent cytotoxic alloimmune responses of natural killer cells (NK) and has been identified as a susceptibility marker predisposing patients to cardiac allograft vasculopathy after heart transplant. This study aimed to investigate whether FCGR2A and FCGR3A polymorphisms can also be associated with the clinical outcome of lung transplant recipients (LTRs). The SNPs of FCGR2A ([131R/H], rs1801274) and FCGR3A ([158V/F], rs396991) were identified in 158 LTRs and 184 Controls (CTL). The corresponding distribution of genotypic and allelic combinations was analyzed for potential links with the development of circulating donor-specific anti-HLA alloantibodies (DSA) detected at months 1 and 3 after lung transplant (LTx), the occurrence of acute rejection (AR) and chronic lung allograft dysfunction (CLAD), and the overall survival of LTRs. The FCGR3A [158V/V] genotype was identified as an independent susceptibility factor associated with higher rates of AR during the first trimester after LTx (HR 4.8, p < 0.0001, 95% CI 2.37-9.61), but it could not be associated with the level of CD16- mediated NK cell activation in response to the LTR's DSA, whatever the MFI intensity and C1q binding profiles of the DSA evaluated. The FCGR2A [131R/R] genotype was associated with lower CLAD-free survival of LTRs, independently of the presence of DSA at 3 months (HR 1.8, p = 0.024, 95% CI 1.08-3.03). Our data indicate that FCGR SNPs differentially affect the clinical outcome of LTRs and may be of use to stratify patients at higher risk of experiencing graft rejection. Furthermore, these data suggest that in the LTx setting, specific mechanisms of humoral alloreactivity, which cannot be solely explained by the complement and CD16-mediated pathogenic effects of DSA, may be involved in the development of acute and chronic lung allograft rejection.

Background: Risk factors for the development of anti-HLA antibodies include blood transfusion, organ transplantation, and pregnancy. Humoral rejection, mediated by donor-specific anti-HLA antibodies (DSA), has been studied in all kind of solid organ transplantations, and several studies have suggested that post-liver transplantation (LT) DSA may play a role in acute and chronic rejection., Objective: The aim of the present study was to assess the impact of pregnancy on the occurrence of DSA and the impact of DSA in a large population of young female LT recipients., Methods: This single center retrospective study included all female patients who underwent a first LT between January 1990 and December 2010 and who were of childbearing age during post-LT follow-up (i.e. 18 to 40 years old)., Results: The study population consisted in 73 patients, and the mean age at LT was 20.9 years (0.6-39.9); 32 patients were transplanted during childhood. The global incidence of de novo DSA was 42.5% (31/73), after a median delay of 15.5 years (1-25) of follow-up after LT. Most de novo DSA were anti-class II alone (90.3%), and included anti-DQ for 80.6%. From the 73 patients, 33 presented at least one pregnancy after LT (45.2%) and before DSA screening. Multivariate analysis disclosed that history of pregnancy (OR = 6.37; 95%CI, 2.17-18.63, p = 0.001) and younger age at LT (OR = 0.96; 95%CI:0.92-0.99, p = 0.033) were significantly associated with de novo DSA. Among the 31 patients who had de novo DSA, the diagnosis of antibody-mediated rejection was made in 8 patients (25.8%), after a median delay of 74 months after LT; 6/8 (75.0%) had history of pregnancy. During follow-up, 3 of these 8 patients lost their liver graft and died., Conclusion: The results of the present study suggest that close monitoring of DSA in young women with history of pregnancy should be recommended regarding the risk of DSA-mediated rejection., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Chronic-active antibody mediated rejection (c-aABMR) contributes significantly to late renal allograft failure. The antibodies directed against donor-derived antigens, e.g. anti-HLA antibodies, cause inflammation at the level of the microvascular endothelium. This is characterized by signs of local activation of the complement system and accumulation of immune cells within the capillaries. Non-invasive biomarkers of c-aABMR are currently not available but could be valuable for early detection. We therefore analyzed the activation profiles of circulating T and B cells, NK cells and monocytes in the peripheral blood of 25 kidney transplant recipients with c-aABMR and compared them to 25 matched recipients to evaluate whether they could serve as a potential biomarker. No significant differences were found in the total percentage and distribution of NK cells, B cells and T cells between the c-aABMRpos and c-aABMRneg cases. There was however a higher percentage of monocytes present in c-aABMRpos cases (p < .05). Additionally, differences were found in activation status of circulating monocytes, NK cells and γδ T cells, mainly concerning the activation marker CD16. Although statistically significant, these differences were not sufficient for use as a biomarker of c-aABMR., (Copyright © 2019 Elsevier B.V. All rights reserved.)

A resurgent interest in the role of B cells following solid organ transplantation is being driven by clinical data suggesting that antibody mediated rejection (AMR) is a major cause of dysfunction and organ transplant failure. These observations suggest that, in a subset of patients, current immunotherapies are failing to control the development of alloantibody responses, and/or failing to reverse the production or the effects of alloantibodies. Quantification of donor-specific antibodies (DSA) has proven to be an imperfect predictor of AMR, and efforts to improve DSA quantification anticipate that this will result in improved predictive power. At the same time, attempts to control of ABMR have focused on the non-specific elimination of B cells, plasma cells (PCs) or circulating antibodies. In the past decade, there has been an improvement in our understanding of the processes that drive B cell differentiation into germinal center (GC)-dependent or GC-independent memory B cells and antibody-secreting PC. These insights are suggesting new ways to more specifically target the DSA response, which may lead to better long-term allograft survival outcomes while preserving protective immunity. In this review, new insights into processes that lead to antibody production upon primary and secondary antigen encounter are discussed, and the potential implications to DSA production as well as future areas of investigation to control AMR are discussed., (Copyright © 2018. Published by Elsevier Inc.)

Background: As CD38 is expressed on red blood cells (RBCs), the plasma of patients on daratumumab (DARA) reacts with the panel cells of pretransfusion tests, masking underlying alloantibodies. The treatment of RBCs with dithiothreitol (DTT) is the most disseminated method to overcome DARA effect on immunohematological tests, but it hampers the identification of potentially harmful antibodies. Our goal was to validate a new strategy, the blockage monoclonal antibody protocol (BMAP), to mitigate the DARA interference on RBCs using anti-CD38 and antihuman globulin., Methods: Samples of patients receiving DARA were included in the study. Sera were tested using both DTT- and BMAP-treated RBCs, which comprised three steps: 1) titration of monoclonal anti-CD38, 2) treatment of RBCs obtained from donors with anti-CD38, and 3) blockage of anti-CD38-adsorbed RBCs with antihuman globulin., Results: Twenty patients were included in the study. Donor RBCs were treated with anti-CD38 and successfully blocked with antihuman globulin. In 19 patients, DARA-mediated agglutination was eliminated using both DTT- and BMAP-treated RBCs. In one patient, agglutination persisted when tested against the BMAP-treated RBCs, and alloantibodies were identified. Patient samples were mixed with commercial anti-D, -C, -e, -K, -Jka, -Kpb and tested against antigen-positive BMAP-treated RBCs, resulting in detection of these antibodies., Conclusion: This study validated a new strategy to minimize the interference of DARA on immunohematological tests. The protocol preserves the integrity of RBC antigens, permitting the detection of antibodies from all blood group systems. The BMAP has potential use in other situations where specific antibodies may interfere with pretransfusion screening., (© 2019 AABB.)

Recent data suggest that HLA epitope matching is more predictive for the formation of donor specific antibodies (DSA) after transplantation than classical HLA antigen matching. A proper definition of the epitopes and their immunogenicity is crucial for a broad clinical application of epitope matching in solid organ transplantation. For that purpose, a novel epitope component was started during the 17th International Histocompatibility and Immunogenetics Workshop. Only a limited number of antibody data was available, resulting in a preliminary analysis confirming the potential relevance of epitope matching for the development of donor specific antibodies. During the actual workshop sessions the participants mainly discussed the future strategy to collect and analyze a large number of data in the framework of the 18th International Histocompatibility and Immunogenetics Workshop, which will take place in 2021. All practical issues can be found on the website www.ihiw18.org. Data collection will already start in early 2019 in order to have sufficient time for detailed analyses using several different algorithms., (Copyright © 2019 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Background: Alloantibodies against more than 50 non-ABO blood group antigens have been implicated in hemolytic disease of the fetus and newborn (HDFN) and are expected to wane within weeks after delivery. Persistent anemia leads to the hypothesis of continued exposure to red blood cell (RBC) alloantibodies via breast milk, which has been shown in a murine model and suggested in rare case reports., Case Report: We report three cases of prolonged HDFN in two neonates with anti-D HDFN and one with anti-Jk a HDFN. Patient 1 demonstrated 4+ anti-D serologic testing beyond 2 months; therefore, antibody testing was performed on maternal breast milk., Methods: Maternal serum samples were tested for the presence of unexpected antibodies using standard Ortho gel card and 37 °C 60 minutes with anti-human globulin (AHG) tube saline methods. Antibody titrations were performed using the standard 37 °C 60 minutes to AHG tube saline method. Fresh breast milk samples were tested using the standard 37 °C 60 minutes to AHG tube saline method for both unexpected antibodies and titration study. Fresh breast milk from an O-positive, antibody-negative donor was used as control for any reactivity that may have been due to milk solids or proteins alone., Results: Using a known methodology applied in a novel way to test breast milk for RBC alloantibodies, antibodies against fetal RBCs were identified in the maternal breast milk of three patients., Conclusion: Maternal RBC alloantibodies are present in breast milk and may be clinically significant in patients with prolonged recovery from HDFN., (© 2019 AABB.)

Chronic antibody-mediated rejection is the leading cause of allograft dysfunction and loss after kidney transplantation, and current immunosuppressive regimens fail to target the plasma cells that produce alloantibodies. We previously showed that treatment with the immunoproteasome inhibitor ONX 0914 prevented the expansion of plasma cells and prevented chronic allograft nephropathy and organ failure after kidney transplantation in rats, but the mechanism has remained elusive. In the current study, we confirmed a long-term reduction in alloantibody production and improvements in allograft histology in rats treated with ONX 0914 or with the broad-spectrum proteasome inhibitor bortezomib. Plasma cells from allotransplanted rats expressed immunoproteasomes at high levels. Immunoproteasome inhibition with ONX 0914 led to ubiquitin-conjugate accumulation, activation of the unfolded protein response, and induction of apoptosis in plasma cells. In addition, ONX 0914 suppressed the expression of adhesion molecules (VLA-4 and LFA-1), plasma cell survival factors (APRIL and IL-6), and IFN-γ-inducible chemokines in bone marrow, while the APRIL receptor BCMA, the IL-6 receptor, and the chemokine receptors CXCR4 and CXCR3 were down-regulated on plasma cells. Taken together, immunoproteasome inhibition blocked alloantibody production by inducing apoptosis of plasma cells through activating the unfolded protein response and suppressing plasma cell survival factors in the bone marrow., (Copyright © 2019 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Our understanding of the conformational and electrostatic determinants that underlie targeting of human leukocyte antigens (HLA) by anti-HLA alloantibodies is principally based upon in silico modelling. Here we provide a biochemical/biophysical and functional characterization of a human monoclonal alloantibody specific for a common HLA type, HLA-A*11:01. We present a 2.4 Å resolution map of the binding interface of this antibody on HLA-A*11:01 and compare the structural determinants with those utilized by T-cell receptor (TCR), killer-cell immunoglobulin-like receptor (KIR) and CD8 on the same molecule. These data provide a mechanistic insight into the paratope-epitope relationship between an alloantibody and its target HLA molecule in a biological context where other immune receptors are concomitantly engaged. This has important implications for our interpretation of serologic binding patterns of anti-HLA antibodies in sensitized individuals and thus, for the biology of human alloresponses.

The International Registry of HLA Epitopes (http://www.epregistry.com.br) is a website-based resource for HLA epitopes important in transplant rejection and platelet transfusion refractoriness. Its primary goal is to document epitopes that are verified experimentally with specific antibodies. Such epitopes can be defined by single eplets and by eplets paired with certain polymorphic residues within a 15-Å radius, the dimension of the corresponding structural epitope. This report is an update of the HLA-ABC repertoire including descriptions of 72 antibody-verifications of epitopes defined by eplets and/or eplet pairs. The newly updated version 2.0 EpRegistry shows also the polymorphic residue compositions of structural epitopes corresponding to eplets shared between groups of alleles. At present, 151 eplets have not been antibody-verified, and we ranked them with a so-called ElliPro score as a potential predictor of immunogenicity. Sixty eplets with low ElliPro scores might be considered non-epitopes incapable of inducing specific antibodies., (Copyright © 2018. Published by Elsevier Inc.)