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Matching for HLA-A, -B, -C, and -DRB1 loci (8/8 match) is currently the gold standard for unrelated donor hematopoietic cell transplantation (HCT). In Europe, patients are also matched at the HLA-DQB1 loci (10/10 match). However, there is increasing evidence that matching at HLA-DRB3/4/5 loci may help to lower transplant-related morbidity and mortality. We therefore investigated the impact of HLA-DRB3/4/5 mismatches on outcomes in 1975 patients who received a first 10/10 matched unrelated donor (MUD) HCT in France from 2000 to 2012 for a hematological malignancy. High-resolution typing was performed at HLA-A, -B, -C, -DRB1, -DQB1, -DPB1, and -DRB3/4/5 loci for all donor/recipient pairs. Compared with DRB3/4/5-matched pairs, patients who received a MUD HCT from a DRB3/4/5 mismatched donor had a significantly increased risk of grade II-IV acute graft-versus-host disease (aGVHD) (Adjusted Hazard Ratio (HR) 1.43 (1.07 to 1.90)) associated with lower graft-versus-host disease-free and relapse-free survival (GRFS) (Adjusted HR 1.20 (1.02 to 1.42)). Conversely, we observed no differences in terms of chronic GVHD, nonrelapse mortality, relapse and overall survival. However, we believe that patients stand to benefit from DRB3/4/5 loci being considered for unrelated donor selection to improve GRFS and then quality of life after unrelated HCT., (© 2018 Wiley Periodicals, Inc.)

Background: Granulocyte antibodies have been implicated in allo- and autoimmune neutropenia and in transfusion reactions., Study Design and Methods: Fifty-one sera from suspected alloimmune neutropenia or transfusion-related acute lung injury (TRALI) and 40 sera from suspected autoimmune neutropenia were tested for granulocyte antibodies using LABScreen MULTI (One Lambda, Inc.), compared with classical tests (flow cytometry [FC] and granulocyte agglutination [GAT] followed by monoclonal antibody-specific immobilization of granulocyte antigens [MAIGA])., Results: In alloimmune situations, 48 sera were concordant (94%), two sera positive for HNA with LABScreen MULTI were negative by FC/GAT and/or MAIGA, and one serum sample negative for HNA with LABScreen MULTI was positive by classical tests. In autoimmune neutropenia, 30 sera were concordant (75%), four sera positive for HNA with LABScreen MULTI were negative by FC/GAT and/or MAIGA, and six sera negative for HNA with LABScreen MULTI were positive by FC/GAT and/or MAIGA. For detection of autoantibodies, the LABScreen MULTI was less concordant. However, with the exception of one case, the discrepancies were observed in sera that did not show a clear specificity., Conclusions: LABScreen MULTI correlated well with our classical methods for HNA-1 and HNA-2a antibody screening. It can be used for screening blood donors or patients suspected of TRALI, but GAT is still needed for HNA-3a antibody screening., (© 2010 American Association of Blood Banks.)

Background/aims: Dendritic cells (DCs), which play a critical role during immune response, could present alternative differentiation patterns depending on tissue microenvironment. Our aim was to examine the influence of hepatic microenvironment on human monocyte differentiation into DCs., Methods: Cytology, immunophenotyping, cytokine production and T-cell activation were analyzed in DCs differentiated from human monocytes co-cultured with rat liver epithelial cells (RLEC) or human cells from various tissue origins and compared to control DCs obtained on plastic with GM-CSF/IL-4., Results: RLEC environment promotes DC differentiation in the presence of IL-4 without GM-CSF. These DCs evidence similar expression of MHC-II, co-stimulatory and adhesion molecules than control DCs, but distinct lineage markers defining a CD11c+/CD14+/CD123+ DC subset. This phenotype is common to DCs from RLEC and human liver environment and differs from that obtained with skin or intestine environments. Functionally, they produce IL-10 but not IL-12p70 and favor IL-4/IL-10 secretion by T-cells rather than IFN-gamma., Conclusions: Our results confirm that tissue niches modulate DC differentiation and demonstrate that hepatic environment influences monocyte differentiation into a DC subset directing Th2 response, a key data for understanding the specialized immune response in liver. They also make RLEC co-culture system useful for studying liver DC functions.

Multiple sclerosis (MS) is a demyelinating autoimmune disease with a strong yet complex genetic component. To date only the HLA-DR locus, and specifically the HLA-DR15 allele, has been identified and confirmed as influencing the risk of developing MS. Genomic screens on several datasets have been performed and have identified several chromosomal regions with interesting results, but none have yet been confirmed. We tested seven of the most-promising regions (on chromosomes 1p, 2p, 3p, 3q, 5q, 19q, and Xp) identified from several genomic screens in a dataset of 98 multiplex MS families from the United States and 90 multiplex MS families from France. The results did not confirm linkage to 2p, 3q, 5q, or Xp in the overall dataset, or in subsets defined by geographic origin or HLA-DR15 status. Regions on 1p34, 3p14, and 19q13 produced lod scores >0.90 in at least one subset of the data, suggesting that these regions should be examined in more detail.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system with a genetic component. Until now, the more consistent association with the disease is found with the major histocompatibility complex, especially HLA-DRB1*1501-DQB1*0602 haplotype. In this report, we demonstrate the interaction of Cytotoxic T Lymphocyte-associated antigen 4 (CTLA-4 [CD152]) gene with DRB1*15 haplotype in multiple sclerosis genetic susceptibility. Our data were obtained from two European independent family-based studies including 610 multiple sclerosis family trios. Ann Neurol 2003;54:119-122

Among candidate genes involved in multiple sclerosis (MS) genetic susceptibility, MHC genes and particularly HLA-DRB1*1501-DQB1*0602 haplotype play a major role. Based on the strong linkage disequilibrium observed in Caucasians between DRB1*1501 and DQB1*0602 alleles, it is still impossible to draw a firm conclusion about the DRB1 or DQB1 locus involvement. In order to address this issue a strategy associating a genetic and a functional approach was conducted in a population of-non-Caucasian MS patients. We observed that in Martinicans (55 MS and 100 controls), the DRB1*15 and DRB1*07 alleles were positively associated with the disease. However in Martinicans the most common DRB1*15 subtype was *1503 and not *1501. Moreover, in Martinicans, the frequency of DQB1*0602, found in association with other DRB1 alleles than DRB1*15 (42% of DQB1*0602 haplotypes), was not increased in DRB1*15-negative MS patients, suggesting a neutral role of DQB1*0602 in MS genetics. In a second step, we demonstrated the capability of the DRB1*1503 allele associated with MS in Martinicans to present the immunodominant autoantigen MBP 85-99 peptide to a DRB1*1501 restricted MBP specific T cell line. Interestingly, structural features of DRB1*1501 or DRB1*1503 molecules are in good fit with the hypothesis that *1501 and *1503 molecules may act similarly in MS development by presenting the same immunodominant MBP peptide. On the whole, our results show a prominent role of the DRB1 locus (DRB1*1501 and/or DRB1*1503 alleles) in the immunodominant MBP 85-99 peptide presentation to genetically different MS patients and suggest a neutral role of the DQB1 encoded molecule in MS susceptibility.

We have developed a real-time quantitative polymerase chain reaction (PCR) assay using TaqMan technology (Applied Biosystems, Foster City, CA) for monitoring donor cell engraftment in allogenic hematopoietic stem cell transplant recipients. For this purpose, we selected 19 specific sequence polymorphisms belonging to 11 human biallelic loci located on 9 different chromosomes. Using a set of specially designed primers and fluorogenic probes, we evaluated the 19 markers' informativity on a panel of 126 DNA samples from 63 recipient/donor pairs. In more than 90% of these pairs, discrimination between recipient and donor genetic profile was possible. By using serial dilutions of mixed DNAs, we evaluated the linearity and sensitivity of the method. A linear correlation with r higher than 0.98 and a sensitivity of 0.1% proved reproducible. Fluorescent-based PCR of short tandem repeats (STR-PCR) and real-time PCR chimerism assay were compared with a panel of artificial cell mixtures. The main advantage of the real-time PCR method over STR-PCR chimerism assays is the absence of PCR competition and plateau biases, and results evidenced greater sensitivity and linearity with the real-time PCR method. Furthermore, different samples can be tested in the same PCR run with a final result in fewer than 48 hours. Finally, we prospectively analyzed patients who received allografts and present 4 different clinical situations that illustrate the informativity level of our method. In conclusion, this new assay provides an accurate quantitative assessment of mixed chimerism that can be useful in guiding early implementation of additional treatments in hematopoietic stem cell transplantation.

The beta subunit of human chorionic gonadotropin (hCG beta) is markedly overexpressed by neoplastic cells of differing histological origin including those present in colon, breast, prostate and bladder tumors. We have previously shown that some patients with hCG beta-producing urothelial tumors have circulating T cells that proliferate in response to hCG beta. To make a comprehensive study of hCG beta as a potential target for cancer immunotherapy, we investigated whether hCG beta peptides could induce CD4+ or CD8+ T-cell responses in vitro. By stimulating peripheral blood mononuclear cells (PBMCs) from three donors with mixtures of overlapping 16-mer synthetic peptides analogous to portions of either the hCG beta 20-71 or the hCG beta 102-129 region, we established six CD4+ T-cell lines that proliferated specifically in response to five distinct determinants located within these two hCG beta regions. Three antigenic determinants (hCG beta 52-67, 106-121 and 114-125) were presented by HLA-DR molecules, while the two other antigenic determinants (hCG beta 48-63 and 56-67) were presented by HLA-DQ molecules. Interestingly, one T-cell line specific for peptide hCG beta 106-121 recognized hCG beta peptides comprising, at position 117, either an alanine or an aspartic acid residue, with the latter residue being present within the protein expressed by some tumor cells. In addition, three other hCG beta-derived peptides that exhibited HLA-A*0201 binding ability were able to stimulate CD8+ cytotoxic T cells from two HLA-A*0201 donors. These three immunogenic peptides corresponded to regions hCG beta 40-48, hCG beta 44-52 and hCG beta 75-84. Our results indicate that the tumor-associated antigen hCG beta possesses numerous antigenic determinants liable to stimulate CD4+ and CD8+ T lymphocytes, and might thus be an effective target antigen for the immunotherapy of hCG beta-producing tumors.

Chemokines and their receptors are involved in the migration of different mononuclear cells. Among them macrophages-derived chemokines (MDC) and thymus-and activation regulated chemokine (TARC) belong to a new cluster of genes involve in Th2 lymphocytes homing. Cytokines appear to play a significant role in pathogenesis of inflammatory bowel diseases with an excessive Th1 response in chronic lesions of Crohn's disease (CD) and a Th2 pattern in both earlier mucosal CD lesions and in mucosa of ulcerative colitis (UC). Here we demonstrate that RNAm coding for MDC and TARC are expressed in mucosa from CD and UC patients. Using real-time fluorescent RT-PCR, MDC and TARC mRNA were increased in CD inflamed mucosa. Moreover MDC and TARC transcripts were increased in inflamed CD specimen compared to non-involved CD mucosa. These differences both discriminate CD from UC patients. Additionally, MDC protein was produced in isolated mononuclear cells from peripheral blood (PBMC) or mucosa (LPMC) from UC and CD patients: spontaneously, MDC production from PBMC was increased in CD compared to UC patients. MDC production from CD PBMC was also higher than that found in healthy controls. Together, these data indicate that MDC should be involved in the lymphocytes homing in mucosa from CD patients.

The Martinican population is mainly the product of admixture between African people and French Caucasians. The aim of the present study is to investigate at the DNA level the polymorphism of HLA class I (HLA-A, HLA-B) and class II (HLA-DRB1, DQB1 and DPB1) genes in a population of 100 Martinicans. Allelic distributions and interlocus linkage disequilibria were compared to those observed in a French Caucasian population and in African or North American African populations. Our data revealed a higher degree of polymorphism in Martinicans than in Caucasians and showed a prominant contribution of African origin in the admixed genetic feature of this population. African characteristic alleles were significantly represented in Martinicans: A*30, *33 *34, *66, *74, *8001, B*1510, *35, *42, *53, DRB1*0302, *0804, *1202, *1304, *1503, DPB1*0101, *1701, *1801, *3901. Moreover a higher diversity of A*-B* and DRB1*-DQB1* associations was observed in Martinicans compared to Caucasians which also reflects the African genetic background of this population. In the whole, using PCR-based genotyping methods for HLA class I and class II loci, this study allows a preliminary description of HLA allele distribution in this Caribbean island and gives new elements which may be helpful in the anthropologic field as well as in HLA and disease association studies.

Objective: HLA DM is a non-classical major histocompatibility complex (MHC) class II molecule that has been shown to facilitate peptide loading with classical class II molecules., Methods: In this study, we analysed the polymorphism in exon 3 of HLA DMA and DMB genes by a polymerase chain reaction-sequence-specific oligonucleotide probe method in 163 rheumatoid arthritis (RA) patients and 146 ethnically matched controls. The HLA-DRB1 genotype was also analysed by a reverse-dot blot method., Results: Our results show in RA patients a significant increase in the HLA DMB*0101 allele frequency (83% vs 72.3% of the controls, P < 1.6 x 10(-3), significance at P < 0.0125) and in the HLA DMB*0101-0101 homozygote genotype frequency [70.8% vs 50% of the controls, P < 4.2 x 10(-4), significance at P < 0.00625, odds ratio (OR) = 2.4, 95% confidence interval (CI): 1.43-4]. The increase in DMB*0101 allele and homozygote genotype frequencies was independent of a linkage disequilibrium between DMB and DRB1 alleles. The analysis of non-random associations between the HLA-DM and DRB1 alleles only revealed a significant association in controls between DMB*0104 and DRB1*07 alleles (delta = 0.01, P < 7 x 10(-4), significance at P < 9.6 x 10(-4)). On the other hand, the DMB*0101-0102 genotype frequency was increased in DRB1*0401-negative RA patients as compared to controls (11% vs 2%, P < 0.011, significance at P < 0.015, OR = 6.2, 95% CI: 1.2-30)., Conclusion: Our data suggest that HLA-DM alleles could play a role in the genetic susceptibility to RA.

In a prospective study between 1993 and 1998, data was collected from 46 pregnant women and subsequently from their babies. 25 pregnant women with active autoimmune thrombocytopenic purpura (AITP) or a history of AITP (group A) and 21 pregnant women with isolated thrombocytopenia and identification of specific platelet autoantibodies detected by monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay (group B) were evaluated for platelet-associated immunoglobulin-G (PAIgG), MAIPA assay and HLA genotype. Neonatal platelet counts were performed at least three times in the first week. 11 neonates were thrombocytopenic (23.9%). No severe haemorrhage occurred. There were no significant differences regarding the values of PAIgG or positive MAIPA tests between mothers of thrombocytopenic or healthy newborns. A significant difference, however, regarding the HLA DRB3* allele was found, with a high incidence in the subgroup of mothers of healthy newborns (P = 0.005). A similar trend was found among mothers with anti-GPIIbIIIa antibodies (P = 0.06). In contrast, HLA DRB5* allele appeared to be present especially in mothers of thrombocytopenic newborns (not significant). Our data suggest that mothers with AITP who have the HLA DRB3* genotype are unlikely to give birth to a thrombocytopenic baby. This study provides a preliminary report on a noninvasive test to identify infants who are likely to be affected.

Objective: Sjögren's syndrome (SS) is a chronic autoimmune disease with a genetic component. Among the genetic factors, the role of HLA class II genes has been suggested and a positive association with DRB1*03 allele has been described. However, there is no consensus on a unique HLA locus for this disease nor on the role of the HLA gene product in the disease. The aim of this study was to analyse prospectively MHC region involvement in the genetic susceptibility to SS by studying DRB1, DQB1, DPB1, TAP1, TAP2 genes and TNF microsatellites in a population of 45 primary SS patients., Methods: All the polymorphisms studied were analysed at the genomic level using PCR-based methodologies., Results: Concerning HLA class II alleles, the highest relative risk to develop the disease was associated with the DRB1*15-DRB1*0301 heterozygous genotype (17.8% vs 3.5% in controls - pc < 0.005, OR = 5.96). Analysing other genes located on the same region allowed us to further determine the DRB1 haplotypes at risk. For instance, the DRB1*0301 haplotype involved in the genetic susceptibility to SS was more often associated with the DPB1* 0201 and TNF-a2 alleles in SS patients than in controls. Moreover, all the DRB1*15-DRB1*0301 SS patients were TAP1-0101, TAP2-0101 homozygous, allowing us to deduce the extended genotype at risk as DRB1*15, TAP1-0101, TAP2-0101/DRB1*0301, TAP1-0101, TAP2-0101 which was carried by only 3 controls out of the 130 tested (p < 0.01, OR = 6.68)., Conclusion: This study confirmed the role of the MHC region in the susceptibility to Sjögren's disease, and for the first time suggests a synergistic interaction between two HLA-DRB1 extended haplotypes in the genetic mechanisms controlling the disease.

Objectives: Multiplex family studies have excluded chromosome 6 as a candidate gene of susceptibility to inflammatory bowel disease. However, one recent study suggested that a gene involved in the pathogenesis of Crohn's disease is located on chromosome 6 confering to a microsatellite allelic combination (a2, b1, c2, d4, e1) a strong genetic risk factor in Crohn's disease. The aim of our study was to determine simultaneously the polymorphisms of the TNF microsatellites and of the genes coding for TNF synthesis in patients with inflammatory bowel disease., Patients and Methods: Sixty patients with ulcerative colitis, 100 patients with Crohn's disease were compared to 64 healthy ethnically matched controls. Five TNF microsatellite loci (a, b, c, d, e) were typed using polymerase chain reaction PCR, and two dimorphisms of TNF alpha and TNF beta (intron 1) were studied by restriction fragment length polymorphism (RFLP)., Results: Allelic frequencies of TNF microsatellites and of TNF alpha and beta genes were similar in Crohn's disease, ulcerative colitis and controls. Five loci microsatellite haplotypes, especially a2 b1 c2 d4 e1 allelic combination, were not more frequent in Crohn's disease (25%) compared to ulcerative colitis (27%) or controls (20%). Subgroups stratification according to clinical characteristics did not modify haplotype frequencies. Analysis of our data taking simultaneously into account the MHC alleles (DRB*01 or DRB1*04) did not modify our data; however, it suggested that extended haplotype on short arm of chromosome 6 differed between patients and controls. Linkage disequilibrium (delta = -360.10(-4); P < 0.01) between a2, b1, c2, d4, e1 allelic combination and DRB1*04 allele was observed only in Crohn's disease., Conclusion: Percentages of patients with Crohn's disease or ulcerative colitis carrying TNF microsatellite or TNF alpha and beta gene haplotypes were similar to those of healthy controls. These data argue against involvement of the TNF locus without exclusion of short arm of chromosome 6 implication in Crohn's disease pathogenesis.

Interaction of the interleukin 2 receptor (IL-2R) beta chain with the lymphocyte-specific protein tyrosine kinase (PTK), p56lck, has led to the speculation that p56lck participates in growth signal transduction. Although activation of T cells with interleukin 2 (IL-2) results in the activation of p56lck, accumulating data support the notion that Lck does not play an essential role in mitogenic signal delivery from the IL-2R. Since this src-related PTK has been shown to enhance TCR/CD3-mediated T cell responsiveness, here we investigated whether activation of Lck by IL-2 could contribute to enhance TCR/CD3-mediated T cell functions. This was achieved by using human CD4(+)-cloned T cells and comparing the effects of IL-2 on p56lck kinase activation and cytokine production. Results show that p56lck kinase activity increased as early as 1 min, reached a maximum within 5 min and decreased within 60 min after IL-2 stimulation. Such treatment with IL-2 also resulted in enhancing T cell responsiveness to CD3+PMA stimulation, as assessed by IL-2 and IFN-gamma secretion and by T cell proliferation. This increase of T cell functions was correlated with IL-2-induced p56lck activation in both dose-response and time-course experiments. Taken together these results strongly suggest that activation of Lck by IL-2 may play a role in regulating CD3-mediated T cell functions.

We show that T cell activation of human CD4+ cloned T cells through the CD2 molecule can induce either autocrine proliferation or cytolysis, depending on the pair of anti-CD2 mAbs used for stimulation, that is, D66/T11(1) or GT2/T11(1), respectively. As the earliest biochemical event after CD2 stimulation is likely the induction of tyrosine phosphorylation of various proteins, we investigated whether differential activation of protein tyrosine kinases (PTKs) could contribute to the selective induction of each function. Results show that herbimycin A, a potent PTK inhibitor, markedly decreased the induction of both proliferation and cytolysis. This implies a regulatory role for tyrosine phosphorylation in the induction of each function by CD2. However, that PTKs are differentially activated upon induction of proliferation by D66/T11(1) or cytotoxic function by GT2/T11(1) emanated from two different approaches. First, immunoblotting total cellular extracts with an anti-phosphotyrosine mAb showed different patterns of tyrosine phosphorylation depending on the pair of CD2 mAbs used for stimulation. Second, a differential activation of p56lck, a src-related PTK, was observed after stimulation with D66/T11, and GT2/T11(1). Although induction of proliferation by D66/T11(1) was correlated with increased Lck activity, this was not observed when cells were triggered to lyse by GT2/T11(1). Thus, by providing striking correlative evidences linking differences in PTK activation with induction of different functions in bifunctional cloned T cells, our results strongly suggest that PTKs may contribute to the selective orientation of T cell functions at a single-cell level.

Two cases of HLA partially matched related cord blood transplantation were reported. The first patient remains healthy more than 4 years after graft. The second patient died from acute GVHD. In the first case, the immune reconstitution was studied over a 2 year period after graft: the alloreactive cytotoxic T-cell precursor (CTLp) frequencies were comparable between the donor and the recipient. Indirect evidence suggested that the resolution of GVHD was not due to a specific deletion of CTLp against the HLA antigens unshared between donor and recipient.