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Philip J Norris - One of the best experts on this subject based on the ideXlab platform.
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c1q binding anti hla antibodies do not predict platelet transfusion failure in trial to reduce alloimmunization to platelets study participants
Transfusion, 2016Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:BACKGROUND In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of 530 subjects became clinically refractory (CR) to platelets (PLTs) without lymphocytotoxicity assay (LCA)-detectable anti-HLA antibodies. The LCA only detects complement-binding antibodies and is less sensitive than newer assays. Utilizing a more sensitive bead-based assay that does not distinguish between complement-binding versus non–complement-binding antibodies, we have previously shown that while many LCA-negative (LCA–) patients do have anti-HLA antibodies, these low- to moderate-level antibodies do not predict refractoriness. As complement can contribute to PLT rejection, we assessed if previously undetected complement-binding antibodies account for refractoriness among LCA– patients. STUDY DESIGN AND METHODS Samples from 169 LCA– (69 CR, 100 non-CR) and 20 LCA-positive (LCA+; 10 CR, 10 non-CR) subjects were selected from the TRAP study serum repository. Anti-Class I HLA immunoglobulin (Ig)G and C1q-binding antibodies were measured in serum or plasma with bead-based detection assays. Levels of C1q-binding antibodies were compared between CR and non-CR subjects and correlated with corrected count increments (CCIs). RESULTS While some of the LCA– subjects had detectable C1q-binding anti-Class I HLA antibodies, and some LCA+ subjects did not, levels were significantly higher among LCA+ subjects. C1q-binding anti-Class I HLA antibody levels did not differ significantly between CR and non-CR among either the LCA– or the LCA+ subjects. Furthermore, there was no significant correlation observed between CCIs and either C1q-binding or any anti-HLA IgG antibodies. CONCLUSIONS This work confirms that low- to moderate-level anti-Class I antibodies do not drive PLT rejection, suggesting a role for antibody-independent mechanisms.
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c1q binding hla antibodies do not predict platelet transfusion failure in trap study participant
Blood, 2014Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:Introduction: In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of the 530 subjects became clinically refractory (CR) to platelet transfusions in the absence of detectable antibodies against HLA as measured by the lymphocytotoxicity assay (LCA). Using more sensitive bead-based detection methods we have previously demonstrated that while many of these LCA- patients do have anti-HLA antibodies, that these low to moderate level antibodies do not predict refractoriness. In addition to being less sensitive then bead based methods, the LCA screen only detects complement-binding antibodies. As these antibodies could be important for platelet rejection, we assessed if previously undetected complement-binding antibodies could account for some of the refractoriness seen in LCA- patients. Methods: 169 LCA- (69 CR, 100 non-CR) and 20 LCA+ (10 CR, 10 non-CR) subjects were selected from the TRAP study. Anti-class I HLA IgG and C1q binding antibodies were measured in serum or plasma using two different multi-analyte, semi-quantitative, bead-based fluorescent antibody detection assays: the LabScreen mixed Luminex assay, and the LabScreen single antigen class I assay with or without added EDTA (One Lambda). Groups were compared using an unpaired t-test, a=0.05, and correlation between variables was also assessed. Results: New measurements of anti-class I HLA IgG antibodies reliably reproduced earlier data with a strong correlation between the old and new measurements (R2=0.9736, p<0.0001). While some of the LCA- subjects did have detectable C1q-binding anti-class I HLA antibodies, and some LCA+ subjects did not, levels of these antibodies were significantly higher among LCA+ subjects (p<0.0001). Levels of C1q-binding anti-class I HLA antibodies were not significantly different between CR and non-CR among either the LCA- or LCA+ subjects. Conclusions: While complement-binding anti-class I HLA antibody levels were higher in the LCA+ subjects, higher levels of these antibodies were not seen in CR LCA+ patients as compared with non-CR LCA+ patients. Complement-binding anti-class I HLA antibodies do not account for refractoriness seen among the LCA- TRAP subjects. This work confirms that low to mid level anti-class I antibodies do not drive refractoriness to platelets, and suggests that antibody-independent mechanisms cause refractoriness in patients lacking higher levels of anti-HLA antibodies. Disclosures No relevant conflicts of interest to declare.
Lisa K. Denzin - One of the best experts on this subject based on the ideXlab platform.
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Inhibition of HLA-DM Mediated MHC Class II Peptide Loading by HLA-DO Promotes Self Tolerance
Frontiers in Immunology, 2013Co-Authors: Lisa K. DenzinAbstract:Major histocompatibility class II (MHCII) molecules are loaded with peptides derived from foreign and self-proteins within the endosomes and lysosomes of antigen presenting cells (APCs). This process is mediated by interaction of MHCII with the conserved, nonpolymorphic MHCII-like molecule HLA-DM (DM). DM activity is directly opposed by HLA-DO (DO), another conserved, non-polymorphic MHCII like molecule. DO is an MHCII substrate mimic. Binding of DO to DM prevents MHCII from binding to DM, thereby inhibiting peptide loading. Inhibition of DM function enables low stability MHC complexes to survive and populate the surface of APCS. As a consequence, DO promotes the display of a broader pool of low abundance self-peptides. Broadening the peptide repertoire theoretically reduces the likelihood of inadvertently acquiring a density of self-ligands that is sufficient to activate self-reactive T cells. One function of DO, therefore, is to promote T cell tolerance by shaping the visible image of self. Recent data also shows that DO influences the adaptive immune response by controlling B cell entry into the germinal center reaction. This review explores the data supporting these concepts.
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HLA-DO increases bacterial superantigen binding to human MHC molecules by inhibiting dissociation of class II-associated invariant chain peptides
Human Immunology, 2013Co-Authors: Abdul Mohammad Pezeshki, Georges A. Azar, Walid Mourad, Jean-pierre Routy, Mohamed Rachid Boulassel, Lisa K. Denzin, Jacques ThibodeauAbstract:Abstract HLA-DO (H2-O in mice) is an intracellular non-classical MHC class II molecule (MHCII). It forms a stable complex with HLA-DM (H2-M in mice) and shapes the MHC class II-associated peptide repertoire. Here, we tested the impact of HLA-DO and H2-O on the binding of superantigens (SAgs), which has been shown previously to be sensitive to the structural nature of the class II-bound peptides. We found that the binding of staphylococcal enterotoxin (SE) A and B, as well as toxic shock syndrome toxin 1 (TSST-1), was similar on the HLA-DO + human B cell lines 721.45 and its HLA-DO − counterpart. However, overexpressing HLA-DO in MHC class II + HeLa cells (HeLa-CIITA-DO) improved binding of SEA and TSST-1. Accordingly, knocking down HLA-DO expression using specific siRNAs decreased SEA and TSST-1 binding. We tested directly the impact of the class II-associated invariant chain peptide (CLIP), which dissociation from MHC class II molecules is inhibited by overexpressed HLA-DO. Loading of synthetic CLIP on HLA-DR + cells increased SEA and TSST-1 binding. Accordingly, knocking down HLA-DM had a similar effect. In mice, H2-O deficiency had no impact on SAgs binding to isolated splenocytes. Altogether, our results demonstrate that the sensitivity of SAgs to the MHCII–associated peptide has physiological basis and that the effect of HLA-DO on SEA and TSST-1 is mediated through the inhibition of CLIP release.
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Sibling rivalry: competition between MHC class II family members inhibits immunity
Nature Structural & Molecular Biology, 2013Co-Authors: Lisa K. Denzin, Peter CresswellAbstract:Peptide loading of major histocompatibility complex (MHC) class II molecules in the endosomes and lysosomes of antigen-presenting cells is catalyzed by human leukocyte antigen-DM (HLA-DM) and modulated by HLA-DO. In a structural study in this issue, Guce et al . show that HLA-DO is an MHC class II mimic and functions as a competitive and essentially irreversible inhibitor of HLA-DM activity, thereby inhibiting MHC class II antigen presentation.
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Evidence for a human leucocyte antigen‐DM‐induced structural change in human leucocyte antigen‐DOβ
Immunology, 2008Co-Authors: Francis Deshaies, Abdul Mohammad Pezeshki, Alexandre Brunet, Angelique Bellemare-pelletier, Djibril A. Diallo, Jean-simon Fortin, Helen M. O’rourke, Nicola Raby, Nathalie Bédard, Lisa K. DenzinAbstract:Human leucocyte antigen (HLA)-DO is a non-classical major histocompatibility complex class II molecule which modulates the function of HLA-DM and the loading of antigenic peptides on molecules such as HLA-DR. The bulk of HLA-DO associates with HLA-DM and this interaction is critical for HLA-DO egress from the endoplasmic reticulum. HLA-DM assists the early steps of HLA-DO maturation presumably through the stabilization of the interactions between the N-terminal regions of the α and β chains. To evaluate a possible role for HLA-DM in influencing the conformation of HLA-DO, we made use of a monoclonal antibody, Mags.DO5, that was raised against HLA-DO/DM complexes. Using transfected cells expressing mismatched heterodimers between HLA-DR and -DO chains, we found that the epitope for Mags.DO5 is located on the DOβ chain and that Mags.DO5 reactivity was increased upon cotransfection with HLA-DM. Our results suggest that HLA-DM influences the folding of HLA-DO in the endoplasmic reticulum. A mutant HLA-DO showing reduced capacity for endoplasmic reticulum egress was better recognized by Mags.DO5 in the presence of HLA-DM. On the other hand, an HLA-DO mutant capable of endoplasmic reticulum egress on its own was efficiently recognized by Mags.DO5, irrespective of the presence of HLA-DM. Taken together, our results suggest that HLA-DM acts as a private chaperone, directly assisting the folding of HLA-DO to promote egress from the endoplasmic reticulum.
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Right place, right time, right peptide: DO keeps DM focused
Immunological Reviews, 2005Co-Authors: Lisa K. Denzin, Jennifer L. Fallas, Maria Prendes, Woelsung YiAbstract:Summary: Peptide loading of major histocompatibility class II molecules is catalyzed in late endosomal and lysosomal compartments of cells by the catalytic action of human leukocyte antigen (HLA)-DM (H-2M in mice). In B cells, dendritic cells and thymic epithelial cells, the peptide loading of class II molecules is modified by the expression of the non-classical class II molecule, HLA-DO (H-2O in mice). Collectively, studies to date support that DO/H-2O expression inhibits the presentation of antigens acquired by cells via fluid phase endocytosis. However, in B cells, the expression of H-2O promotes the presentation of antigens internalized by the B-cell receptor. In this review, we summarize the literature pertaining to DO assembly, transport, and function, with an emphasis on the function of DO/H-2O.
Sherrill J Slichter - One of the best experts on this subject based on the ideXlab platform.
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c1q binding anti hla antibodies do not predict platelet transfusion failure in trial to reduce alloimmunization to platelets study participants
Transfusion, 2016Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:BACKGROUND In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of 530 subjects became clinically refractory (CR) to platelets (PLTs) without lymphocytotoxicity assay (LCA)-detectable anti-HLA antibodies. The LCA only detects complement-binding antibodies and is less sensitive than newer assays. Utilizing a more sensitive bead-based assay that does not distinguish between complement-binding versus non–complement-binding antibodies, we have previously shown that while many LCA-negative (LCA–) patients do have anti-HLA antibodies, these low- to moderate-level antibodies do not predict refractoriness. As complement can contribute to PLT rejection, we assessed if previously undetected complement-binding antibodies account for refractoriness among LCA– patients. STUDY DESIGN AND METHODS Samples from 169 LCA– (69 CR, 100 non-CR) and 20 LCA-positive (LCA+; 10 CR, 10 non-CR) subjects were selected from the TRAP study serum repository. Anti-Class I HLA immunoglobulin (Ig)G and C1q-binding antibodies were measured in serum or plasma with bead-based detection assays. Levels of C1q-binding antibodies were compared between CR and non-CR subjects and correlated with corrected count increments (CCIs). RESULTS While some of the LCA– subjects had detectable C1q-binding anti-Class I HLA antibodies, and some LCA+ subjects did not, levels were significantly higher among LCA+ subjects. C1q-binding anti-Class I HLA antibody levels did not differ significantly between CR and non-CR among either the LCA– or the LCA+ subjects. Furthermore, there was no significant correlation observed between CCIs and either C1q-binding or any anti-HLA IgG antibodies. CONCLUSIONS This work confirms that low- to moderate-level anti-Class I antibodies do not drive PLT rejection, suggesting a role for antibody-independent mechanisms.
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c1q binding hla antibodies do not predict platelet transfusion failure in trap study participant
Blood, 2014Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:Introduction: In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of the 530 subjects became clinically refractory (CR) to platelet transfusions in the absence of detectable antibodies against HLA as measured by the lymphocytotoxicity assay (LCA). Using more sensitive bead-based detection methods we have previously demonstrated that while many of these LCA- patients do have anti-HLA antibodies, that these low to moderate level antibodies do not predict refractoriness. In addition to being less sensitive then bead based methods, the LCA screen only detects complement-binding antibodies. As these antibodies could be important for platelet rejection, we assessed if previously undetected complement-binding antibodies could account for some of the refractoriness seen in LCA- patients. Methods: 169 LCA- (69 CR, 100 non-CR) and 20 LCA+ (10 CR, 10 non-CR) subjects were selected from the TRAP study. Anti-class I HLA IgG and C1q binding antibodies were measured in serum or plasma using two different multi-analyte, semi-quantitative, bead-based fluorescent antibody detection assays: the LabScreen mixed Luminex assay, and the LabScreen single antigen class I assay with or without added EDTA (One Lambda). Groups were compared using an unpaired t-test, a=0.05, and correlation between variables was also assessed. Results: New measurements of anti-class I HLA IgG antibodies reliably reproduced earlier data with a strong correlation between the old and new measurements (R2=0.9736, p<0.0001). While some of the LCA- subjects did have detectable C1q-binding anti-class I HLA antibodies, and some LCA+ subjects did not, levels of these antibodies were significantly higher among LCA+ subjects (p<0.0001). Levels of C1q-binding anti-class I HLA antibodies were not significantly different between CR and non-CR among either the LCA- or LCA+ subjects. Conclusions: While complement-binding anti-class I HLA antibody levels were higher in the LCA+ subjects, higher levels of these antibodies were not seen in CR LCA+ patients as compared with non-CR LCA+ patients. Complement-binding anti-class I HLA antibodies do not account for refractoriness seen among the LCA- TRAP subjects. This work confirms that low to mid level anti-class I antibodies do not drive refractoriness to platelets, and suggests that antibody-independent mechanisms cause refractoriness in patients lacking higher levels of anti-HLA antibodies. Disclosures No relevant conflicts of interest to declare.
Jacques Thibodeau - One of the best experts on this subject based on the ideXlab platform.
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HLA-DO increases bacterial superantigen binding to human MHC molecules by inhibiting dissociation of class II-associated invariant chain peptides
Human Immunology, 2013Co-Authors: Abdul Mohammad Pezeshki, Georges A. Azar, Walid Mourad, Jean-pierre Routy, Mohamed Rachid Boulassel, Lisa K. Denzin, Jacques ThibodeauAbstract:Abstract HLA-DO (H2-O in mice) is an intracellular non-classical MHC class II molecule (MHCII). It forms a stable complex with HLA-DM (H2-M in mice) and shapes the MHC class II-associated peptide repertoire. Here, we tested the impact of HLA-DO and H2-O on the binding of superantigens (SAgs), which has been shown previously to be sensitive to the structural nature of the class II-bound peptides. We found that the binding of staphylococcal enterotoxin (SE) A and B, as well as toxic shock syndrome toxin 1 (TSST-1), was similar on the HLA-DO + human B cell lines 721.45 and its HLA-DO − counterpart. However, overexpressing HLA-DO in MHC class II + HeLa cells (HeLa-CIITA-DO) improved binding of SEA and TSST-1. Accordingly, knocking down HLA-DO expression using specific siRNAs decreased SEA and TSST-1 binding. We tested directly the impact of the class II-associated invariant chain peptide (CLIP), which dissociation from MHC class II molecules is inhibited by overexpressed HLA-DO. Loading of synthetic CLIP on HLA-DR + cells increased SEA and TSST-1 binding. Accordingly, knocking down HLA-DM had a similar effect. In mice, H2-O deficiency had no impact on SAgs binding to isolated splenocytes. Altogether, our results demonstrate that the sensitivity of SAgs to the MHCII–associated peptide has physiological basis and that the effect of HLA-DO on SEA and TSST-1 is mediated through the inhibition of CLIP release.
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Cutting Edge: HLA-DO Impairs the Incorporation of HLA-DM into Exosomes
Journal of Immunology, 2011Co-Authors: Marie-hélène Côté, Marie-claude Bourgeois-daigneault, Alexandre Brunet, Marie-Élaine Gauvreau, Andrew D. Shaw, Jacques ThibodeauAbstract:In multivesicular bodies, HLA-DM (DM) assists the loading of antigenic peptides on classical MHC class II molecules such as HLA-DR. In cells expressing HLA-DO (DO), DM is redistributed from the internal vesicles to the limiting membrane of these organelles. This suggests that DO might reduce DM incorporation into exosomes, which are shed upon fusion of multivesicular bodies with the plasma membrane. To test this hypothesis, we used the 721.45 B lymphoblastoid cell line and different HeLa cell transfectants. We demonstrate that the poor recovery of DM in exosomes as compared with HLA-DR is not the mere reflection of differences in protein expression. Indeed, we found that DO contributes to the inefficient transfer of DM to exosomes. This negative regulation requires an intact di-leucine endosomal sorting motif in the cytoplasmic tail of HLA-DOβ. These results demonstrate that canonical sorting signals and protein–protein interactions modulate the selection of MHC protein cargos.
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A point mutation in the groove of HLA-DO allows egress from the endoplasmic reticulum independent of HLA-DM
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Francis Deshaies, Lisa K. Denzin, Angela Samaan, Alexandre Brunet, Djibril A. Diallo, Jacques ThibodeauAbstract:B lymphocytes express the nonclassical class II molecule HLA-DO, which modulates the peptide loading activity of HLA-DM in the endocytic pathway. Binding to HLA-DM is required for HLA-DO to egress from the endoplasmic reticulum (ER). To gain insights into the mode of action of DO and on the role of DM in ER release, we sought to identify DM-binding residues on DO. Our results show that DOα encompasses the binding site for HLA-DM. More specifically, mutation of residue DOα41 on an exposed lateral loop of the α1 domain affects the binding to DM, ER egress, and activity of DO. Using a series of chimeric DR/DO molecules, we confirmed the role of the α chain and established that a second DM-binding region is located C-terminal to the DOα80 residue, most probably in the α2 domain. Interestingly, after mutation of a buried proline (α11) on the floor of the putative peptide-binding groove, HLA-DO remained functional but became independent of HLA-DM for ER egress and intracellular trafficking. Collectively, these results suggest that the binding of HLA-DM to DOα allows the complex to egress from the ER by stabilizing intramolecular contacts between the N-terminal antiparallel β-strands of the DOαβ heterodimer.
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HLA-DO transduced in human monocyte-derived dendritic cells modulates MHC class II antigen processing
Journal of Leukocyte Biology, 2005Co-Authors: Angelique Bellemare-pelletier, Jean-pierre Routy, Mohamed Rachid Boulassel, Jessy Tremblay, Sylvie Beaulieu, Bernard Massie, Réjean Lapointe, Jacques ThibodeauAbstract:Through the regulation of human leuko- cyte antigen (HLA)-DM (DM) in B cells, HLA-DO (DO) modulates positively or negatively the presen- tation of specific peptides. Transduction of DO into human blood monocyte-derived dendritic cells (MoDC) has been proposed as a mean of modifying the peptide repertoire of major histocompatibility complex class II molecules. However, maturation of DC induced by inflammatory stimuli or possibly the adenoviral vector itself triggers acidification of vesi- cles and shuts down transcription of the class II trans- activator gene as well as de novo biosynthesis of class II-related molecules and DM activity. In these condi- tions, it is unclear that transduced DO could alter the peptide repertoire. Our Western blot and reverse transcriptase-polymerase chain reaction analyses re- vealed that human DC derived from blood monocytes express small amounts of DO. Transduction of DO alone resulted in the accumulation of a small pool of DO in DMCD63vesicles and at the plasma membrane of mature DC. The cell-surface increase in class II-associated invariant chain peptide (CLIP)/ class II complexes is in line with an inhibitory role of DO on DM. Cotransduction of DO and DO only slightly increased CLIP and DO levels at the cell surface. Together with the fact that a large fraction of transduced DO remains in the endoplasmic reticu- lum, this suggests that DM is limiting in these condi- tions. DO expression did not affect a mixed lympho- cyte reaction but reduced presentation of the exog- enous gp100 antigen to a specific T cell clone. These results show that transduced DO modulates antigen presentation in human mature MoDC, evoking the possible use of this chaperone for immunotherapy. J. Leukoc. Biol. 78: 95-105; 2005.
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Class II transactivator-induced expression of HLA-DOβ in HeLa cells
Tissue Antigens, 2002Co-Authors: Hayssam Khalil, Alexandre Brunet, Francis Deshaies, Angelique Bellemare-pelletier, A. Faubert, G.a. Azar, Jacques ThibodeauAbstract:: HLA-DO is an intracellular nonclassical MHC class II molecule expressed in the endocytic pathway of B lymphocytes. It shapes the repertoire of peptides bound to classical class II molecules such as HLA-DR by regulating the activity of HLA-DM. Using a peptide corresponding to the cytoplasmic tail of HLA-DOβ, we have developed a mouse monoclonal antibody, HKC5. Immunofluorescence microscopy revealed that HKC5 recognizes HLA-DO molecules present in the endoplasmic reticulum as well as those in vesicular compartments of the endocytic pathway. In addition, the antibody detects the isolated β chain on Western blots. Using mutants of the DOβ cytoplasmic tail fused to a reporter molecule and expressed in epithelial cells, we showed by flow cytometry that the antibody epitope includes one or both of the leucine residues forming the lysosomal sorting signal. Finally, we have used HKC5 to evaluate the presence of the HLA-DOβ chain in HeLa cells expressing the class II transactivator protein CIITA. Our flow cytometry and confocal microscopy analyses showed a marked expression of DOβ suggesting that HLA-DO could accumulate under the influence of CIITA in non-B cells.
Rachael P Jackman - One of the best experts on this subject based on the ideXlab platform.
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c1q binding anti hla antibodies do not predict platelet transfusion failure in trial to reduce alloimmunization to platelets study participants
Transfusion, 2016Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:BACKGROUND In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of 530 subjects became clinically refractory (CR) to platelets (PLTs) without lymphocytotoxicity assay (LCA)-detectable anti-HLA antibodies. The LCA only detects complement-binding antibodies and is less sensitive than newer assays. Utilizing a more sensitive bead-based assay that does not distinguish between complement-binding versus non–complement-binding antibodies, we have previously shown that while many LCA-negative (LCA–) patients do have anti-HLA antibodies, these low- to moderate-level antibodies do not predict refractoriness. As complement can contribute to PLT rejection, we assessed if previously undetected complement-binding antibodies account for refractoriness among LCA– patients. STUDY DESIGN AND METHODS Samples from 169 LCA– (69 CR, 100 non-CR) and 20 LCA-positive (LCA+; 10 CR, 10 non-CR) subjects were selected from the TRAP study serum repository. Anti-Class I HLA immunoglobulin (Ig)G and C1q-binding antibodies were measured in serum or plasma with bead-based detection assays. Levels of C1q-binding antibodies were compared between CR and non-CR subjects and correlated with corrected count increments (CCIs). RESULTS While some of the LCA– subjects had detectable C1q-binding anti-Class I HLA antibodies, and some LCA+ subjects did not, levels were significantly higher among LCA+ subjects. C1q-binding anti-Class I HLA antibody levels did not differ significantly between CR and non-CR among either the LCA– or the LCA+ subjects. Furthermore, there was no significant correlation observed between CCIs and either C1q-binding or any anti-HLA IgG antibodies. CONCLUSIONS This work confirms that low- to moderate-level anti-Class I antibodies do not drive PLT rejection, suggesting a role for antibody-independent mechanisms.
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c1q binding hla antibodies do not predict platelet transfusion failure in trap study participant
Blood, 2014Co-Authors: Rachael P Jackman, Douglas Bolgiano, Mila Lebedeva, Sherrill J Slichter, Philip J NorrisAbstract:Introduction: In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of the 530 subjects became clinically refractory (CR) to platelet transfusions in the absence of detectable antibodies against HLA as measured by the lymphocytotoxicity assay (LCA). Using more sensitive bead-based detection methods we have previously demonstrated that while many of these LCA- patients do have anti-HLA antibodies, that these low to moderate level antibodies do not predict refractoriness. In addition to being less sensitive then bead based methods, the LCA screen only detects complement-binding antibodies. As these antibodies could be important for platelet rejection, we assessed if previously undetected complement-binding antibodies could account for some of the refractoriness seen in LCA- patients. Methods: 169 LCA- (69 CR, 100 non-CR) and 20 LCA+ (10 CR, 10 non-CR) subjects were selected from the TRAP study. Anti-class I HLA IgG and C1q binding antibodies were measured in serum or plasma using two different multi-analyte, semi-quantitative, bead-based fluorescent antibody detection assays: the LabScreen mixed Luminex assay, and the LabScreen single antigen class I assay with or without added EDTA (One Lambda). Groups were compared using an unpaired t-test, a=0.05, and correlation between variables was also assessed. Results: New measurements of anti-class I HLA IgG antibodies reliably reproduced earlier data with a strong correlation between the old and new measurements (R2=0.9736, p<0.0001). While some of the LCA- subjects did have detectable C1q-binding anti-class I HLA antibodies, and some LCA+ subjects did not, levels of these antibodies were significantly higher among LCA+ subjects (p<0.0001). Levels of C1q-binding anti-class I HLA antibodies were not significantly different between CR and non-CR among either the LCA- or LCA+ subjects. Conclusions: While complement-binding anti-class I HLA antibody levels were higher in the LCA+ subjects, higher levels of these antibodies were not seen in CR LCA+ patients as compared with non-CR LCA+ patients. Complement-binding anti-class I HLA antibodies do not account for refractoriness seen among the LCA- TRAP subjects. This work confirms that low to mid level anti-class I antibodies do not drive refractoriness to platelets, and suggests that antibody-independent mechanisms cause refractoriness in patients lacking higher levels of anti-HLA antibodies. Disclosures No relevant conflicts of interest to declare.