Biomedical subjects

E M Mickelson

Publications and source records attributed to E M Mickelson.

At least 37 records · Page 2Linked to original sources

PubMed1992

Mixed leukocyte culture reactivity and graft-versus-host disease in HLA-identical marrow transplantation for leukemia.

The results of pretransplant mixed leukocyte culture (MLC) assays were compared to subsequent risk of graft-versus-host disease (GVHD) in 783 patients receiving marrow transplants from HLA genotypically identical sibling donors. The mean MLC response observed between 1303 normal HLA identical sibling pairs was 0.0 +/- 4.2% RR. The donor anti-recipient MLC reaction, an in vitro response that presumably might be relevant to GVHD, was significantly increased (greater than mean + 2 sd) in 83 (10.6%) of the cases, most often in patients in relapse at the time of testing. No association was found, however, between this increased donor anti-recipient MLC reactivity pretransplant and the incidence or severity of subsequent acute or chronic GVHD. These data suggest that the increased MLC responses sometimes observed between leukemia patients and their HLA identical sibling donors prior to marrow transplantation do not represent genetic differences capable of causing GVHD.

Bone Marrow Transplantation↗

PubMed1991

Ten HLA-DR4 alleles defined by sequence polymorphisms within the DRB1 first domain.

We have studied DRB1 sequence polymorphisms associated with DR4 subtypes using DR4-specific DNA amplification and sequence-specific oligonucleotide probe (SSOP) hybridization. The 5' amplification primer was designed to hybridize with a unique sequence in the first hypervariable region (HVR) of the DRB1 second exon of all known DR4 alleles; the 3' primer was designed to hybridize with an intron sequence common to all DRB1 The specificity of the amplification step was demonstrated by double-blind testing of 105 selected DNA samples. Prospective SSOP typing of DR4 alleles was performed in 104 unrelated individuals known to be DR4-positive, including 13 who were DR4-homozygous. A DRB1 subtype corresponding with the previously defined DR4-associated specificities Dw4, Dw10, Dw13.1, Dw13.2, Dw14.1, Dw14.2, Dw15, and DwKT2 could be assigned for each of the 117 DR4 haplotypes tested. In most cases, DR4-homozygous, DRB1-heterozygous individuals could be genotyped with the panel of probes. In the course of our analysis, we identified two new DR4-related alleles, DRB1*04.CB (DRB1*0410)1 and DRB1*04.EC (DRB1*, 0411)2 which were recognized by their novel hybridization patterns. The DRB1 second exon sequence of DRB1*04.CB is identical to DRB1*0405 except at codon 86 where GTG encodes valine instead of GGT encoding glycine. DRB1*04.EC is identical to DRB1*04.CB except at codon 74 where GAG encodes glutamic acid instead of GCG encoding alanine. Our results provide further evidence that SSOP hybridization is the most effective approach available for subtyping DR4 haplotypes and identifying unrecognized variants. A similar approach should be equally informative for subtyping other DR alleles.

Alleles↗

PubMed1991

T-cell clones identify three distinct epitopes associated with HLA-Dw14.

Alloreactive T-cell clones were used to study allodeterminants associated with the HLA-DR1, -DR4, and -DRw14 allelic families. Three clones derived by priming against the DR4,Dw14 alloantigen were tested against a panel of HLA-D homozygous B-cell lines and homozygous and heterozygous peripheral blood lymphocytes. Each clone was blocked by monoclonal antibodies specific for HLA-DR, but not HLA-DQ or -DP, molecules, and each showed a unique pattern of allorecognition when tested against the cell panel. Clone 14B appeared to recognize a specific sequence, termed L67-A74, comprised of amino acids in the third hypervariable region of the alpha-helix of the DR beta 1 molecule, and expressed on certain DR1-, DR4-, and DRw14-positive cells. Clone EMO25 recognized the same L67-A74 sequence, but only when expressed on DR4-positive cells, suggesting a role for residues in the first and second hypervariable regions of DR4-positive DR beta 1 molecules in T-cell recognition. Clone EM036 also recognized the L67-A74 sequence, but only when expressed on DR4,Dw14.1-positive cells, implicating residues at positions 57 and 86 of the alpha-helix in T-cell recognition. These results demonstrate the range of specific T-cell responses that are possible against alloepitopes expressed by a single class II allele (Dw14), and are an indication of the diverse regions of the class II molecule that can contribute to allorecognition sites.

Amino Acid Sequence↗

PubMed1991

Alloreactive T-cell clones identify multiple HLA-DQw3 variants.

HLA-DQw3 is a broadly defined alloantigen that has been subdivided by serological, biochemical, and molecular methods into three distinct specificities: DQw7, DQw8, and DQw9. In order to characterize functionally relevant structural polymorphisms within this family of alloantigens, we generated a series of DQw3-reactive T-cell clones that together recognize six different variants of DQw3. T-cell clones IG11 and IG9 were found to recognize three distinct functional variants associated with a majority of DQw3+ cells, while clones 21J, IE6, 64B, and IC3 recognized four more narrowly distributed functional variants associated with unique DQw7, DQw8, and DQw9 subsets. Comparison of known DQB gene sequences suggested candidate recognition sites for clones IG11 and 64B in the region of amino acid residues 66 to 71 and residue 57 of the DQ beta chain. In contrast, no unique DQB or DQA sequences were found that individually corresponded to the reactivity patterns of clones 21J, IE6, IG9, or IC3, suggesting that an interaction between DQ alpha and DQ beta chains determines allo-recognition. These data are consistent with the hypothesis that T cells recognize specific alloepitopes on HLA class II molecules, either as distinct structural elements that trigger an alloresponse or, more indirectly, as contact elements that influence alloreactivity by governing the binding of foreign peptide. The results illustrate the diversity of possible T cell responses directed toward HLA-DQ molecules and suggest that T cell recognition of the DQ heterodimer alone, or a peptide antigen bound to the DQ heterodimer, can be affected either by the individual DQ alpha and beta chains, or by a more complex interaction between the two.

Alleles↗

PubMed1991

Marrow transplantation from HLA-matched unrelated donors for treatment of hematologic malignancies.

Less than 40% of the patients who could benefit from marrow transplantation have an HLA-matched relative who can serve as a donor. For this reason, several centers have explored marrow transplantation from other categories of donors. This retrospective study analyzes the results of marrow transplantation for 52 patients receiving grafts from HLA-A,B,DR,Dw-phenotypically matched, MLC-compatible, unrelated volunteer donors compared to a disease, disease-stage, and age-matched cohort of 104 patients transplanted from HLA-genotypically identical sibling donors. The patients transplanted from unrelated donors had an increased incidence of grade II-IV acute graft-versus-host disease compared to patients transplanted from related donors (79% vs. 36%, P much less than 0.001). However, the probability of relapse-free survival appears similar in the two groups (P = 0.39 over all, with estimates of 41% vs. 46% at 1 year). We conclude from this preliminary data that marrow transplantation from HLA-matched unrelated donors should be considered in most, if not all, circumstances where transplantation from an HLA-matched sibling would be indicated if such a donor were available.

Acute Disease↗

PubMed1991

Transfusions shortly before HLA-matched marrow transplantation for leukemia are associated with a decrease in chronic graft-versus-host disease.

The effect of random red cell transfusions given shortly before allogeneic bone marrow transplantation (BMT) was evaluated in 969 leukemic patients transplanted from an HLA-identical sibling donor. Patients were divided into two groups: 501 who received a transfusion shortly before BMT, and 468 who did not. Both groups had a similar incidence of acute graft-versus-host disease (GVHD), but the recently-transfused group had a significantly lower incidence of chronic GVHD (35.9% vs 48.9%). These differences remained significant in a multivariate analysis of time to chronic GVHD (p = 0.022), taking into account other differences between the two groups and known risk factors for chronic GVHD.

Adult↗

PubMed1990

Mapping of distinct serologic and T cell recognition epitopes on an HLA-DR beta-chain.

A new DR beta-chain allele is defined that is identical to the previously described DR6b molecule except for the first hyperpolymorphic region, where the new allele displays the same polymorphisms found on DR8 and DR12 genes. Two distinct epitopes have been mapped on this new allele. The polymorphism in common with DRw8 and DRw12 is recognized by mAb GS313-9D11. However, alloreactive T cell clones specific for DR6b cells (Dw9) recognize this allele, whereas Dw8-specific T cell clones do not. The mAb determinant maps to the first beta-sheet and probably involves a polymorphic residue lying outside the helix. The binding of mAb 9D11 to this region does not interfere with TCR binding. Alloreactive T cell recognition is associated with polymorphisms located predominantly on the alpha-helical portion of the molecule.

Alleles↗

PubMed1990

Unique sequences for two HLA-DRB1 genes expressed on distinct DRw6 haplotypes.

We have used restriction fragment length polymorphism (RFLP) analysis and DNA sequencing to characterize two distinct DRB1 alleles expressed on DRw52 and DQw7-associated haplotypes but not readily defined by conventional DR serology. These two haplotypes, designated HLA-D "HAG" and "PEV", react variably with DRw13(w6), DRw14(w6), and the more broad DR"3 + 6" antisera. Analysis of RFLP revealed that HLA-D "HAG" and "PEV" are associated with different DRw52 variants, and that "HAG" is indistinguishable from DRw18(3) haplotypes. Sequencing of the "HAG" and "PEV" DRB1 genes showed each to represent novel alleles. Nevertheless, these sequences show similarities with the other alleles of the DR5,w6, and w8 family. "HAG" (DRB1*1303) appears to have arisen either from two recombinational events involving at least three DRB1 sequences (DRB1*1101, DRB1*0803, DRB1*0401) or from a single recombinational event together with multiple point mutational events. "PEV" appears to represent a DRB1*1301-1302/DRB1*1101 recombinant allele, with recombination having occurred in the region of bases 175-198. The results of this study suggest that the DRw52 family of haplotypes is derived from a relatively restricted number of ancestral sequences, with diversity among DRB1 alleles within this family arising through gene conversion or recombination events.

Amino Acid Sequence↗

PubMed1990

Structural requirements for recognition of the HLA-Dw14 class II epitope: a key HLA determinant associated with rheumatoid arthritis.

Although HLA genes have been shown to be associated with certain diseases, the basis for this association is unknown. Recent studies, however, have documented patterns of nucleotide sequence variation among some HLA genes associated with a particular disease. For rheumatoid arthritis, HLA genes in most patients have a shared nucleotide sequence encoding a key structural element of an HLA class II polypeptide; this sequence element is critical for the interaction of the HLA molecule with antigenic peptides and with responding T cells, suggestive of a direct role for this sequence element in disease susceptibility. We describe the serological and cellular immunologic characteristics encoded by this rheumatoid arthritis-associated sequence element. Site-directed mutagenesis of the DRB1 gene was used to define amino acids critical for antibody and T-cell recognition of this structural element, focusing on residues that distinguish the rheumatoid arthritis-associated alleles Dw4 and Dw14 from a closely related allele, Dw10, not associated with disease. Both the gain and loss of rheumatoid arthritis-associated epitopes were highly dependent on three residues within a discrete domain of the HLA-DR molecule. Recognition was most strongly influenced by the following amino acids (in order): 70 greater than 71 greater than 67. Some alloreactive T-cell clones were also influenced by amino acid variation in portions of the DR molecule lying outside the shared sequence element.

Antibodies, Monoclonal↗

PubMed1989

Correlation of structure with T cell responses of the three members of the HLA-DRw52 allelic series.

A third allele at the DRB3 locus, DRw52c, represents an intermediate sequence between DRw52a and DRw52b and may have arisen by a gene conversion-like event. The recognition of cells bearing these molecules by a number of alloreactive and antigen-specific DR-restricted T cell clones was analyzed. On the basis of a theoretical model of HLA class II structure, distinct amino acid clusters have been identified as motifs controlling TCR recognition. These are located both in the cleft and in the alpha-helical edge of the MHC class II recognition platform. Motifs shared between two alleles may restrict public T cell clones.

Alleles↗

PubMed1988

Variants of HLA-DRw52 defined by T lymphocyte clones.

Polymorphism within the gene encoding the DRw52 allospecificity was studied with DRw52-specific proliferative T lymphocyte clones. Three clones, C6, E3 and ZUK16, were generated by intra-DRw52 priming in mixed lymphocyte culture and tested against an HLA-D homozygous reference cell panel. The reactivity of each clone could be specifically inhibited by anti-DR, but not anti-DQ or anti-DP, monoclonal antibodies. Clone C6 identified a DRw52 variant termed 52a that was predominantly expressed by HLA-B8,DR3+ and DRw13,Dw18+ cells. Clone E3 identified a variant termed 52b which was predominantly expressed by HLA-B18,DR3+,DRw11,Dw5+ and DRw14,Dw9+ cells. Clone ZUK16 identified a variant termed 52c which was predominantly expressed by DRw13,Dw19+ cells. The DRw52a, 52b and 52c variants correspond to the Dw24, Dw25 and Dw26 alleles defined by the WHO HLA 1987 Nomenclature Committee. Together, clones E3 and ZUK16 appeared to identify a fourth DRw52 variant termed 52d which was expressed by two cells, one DR3, Dw"3.3"+ and one DRw14,Dw"9.2"+. A fifth Drw52 variant termed 52e, expressed by a DRw13,Dw"OMW"+ cell, was suggested by the absence of reactivity with any of the three T cell clones. These data thus demonstrate the existence of three well-defined allelic variants of DRw52 and indicate that there are at least two additional variants. The recognition of these polymorphisms by alloreactive T cells provides one measure of their functional significance.

Alleles↗

PubMed1988

T-cell-defined DR4 subtypes as markers for type 1 diabetes.

In most populations studied, HLA-DR4, a DRB1 (formerly DR beta I) allele, is increased in frequency among patients with insulin-dependent diabetes mellitus (IDDM). Using T-cells, one can distinguish five subtypes of DR4 (Dw4, Dw10, Dw13, Dw14, and Dw15). Two of these (Dw4 and Dw10) are IDDM-associated in the populations studied here. Therefore, Dw4 and Dw10 could be causative or merely markers for a linked diabetes allele. If they are causative, one might expect them to share some unique structural element or at least to associate consistently with IDDM in different populations. Published sequence data show no structural element unique to Dw4 and Dw10; moreover, the associations of these DR4-Dw subtypes with diabetes vary considerably in different populations. Thus the DRB1 locus probably cannot account for the DR4 association in IDDM. The strong linkage disequilibrium between IDDM and Dw4 and Dw10 suggests that the diabetes susceptibility locus should be in the vicinity of the DR region or the DQ region of the HLA complex. Alternative hypotheses are discussed, relating DR- and DQ-region alleles to IDDM. We further postulate that the evolutionary event that produced the Dw10 allele occurred on a Dw4 haplotype that happened to carry a diabetes susceptibility allele at another locus.

Adolescent↗

PubMed1988

DQw3 variants defined by cloned alloreactive T cells.

The polymorphism of HLA class II molecules expressing the serologically defined alloantigen DQw3 was studied using cloned proliferative T lymphocytes. Two clones, IG9 and IC3, were selectively primed against DQw3-associated determinants and tested against a panel of 92 HLA-D homozygous cells. Both clones were specific for DQw3, but each showed a distinct response pattern. Clone IG9 recognized a DQw3-associated determinant expressed on a subset of DR4 and DR5 haplotypes and on all DRw6, 7, w8, and w9 haplotypes tested. In contrast, clone IC3 recognized a distinct DQw3-associated determinant expressed only on a subset of DR4 haplotypes. In monoclonal antibody inhibition experiments, anti-DQ, but not anti-DR or anti-DP antibodies, blocked reactivity of both clones IG9 and IC3, further demonstrating that the determinants defined by these clones are associated with DQ molecules. In DNA hybridization studies using a DQ beta probe, a correlation was observed between restriction site polymorphisms in the DQ beta gene, designated DQw"3.1" and "3.2," and the expression of the T-cell-defined IG9 and IC3 determinants. It is, thus, possible to demonstrate by cloned T-cell reactivity functionally relevant recognition sites on DQw3+ molecules that are associated with structural polymorphisms defined by molecular and genomic analysis.

Antibodies, Monoclonal↗

PubMed1988

Probability of finding HLA-matched unrelated marrow donors.

Bone marrow transplantation has become the treatment of choice for certain hematologic diseases. However, only 30-40% of patients who might benefit from this procedure have a suitable family donor. Consequently, many centers have begun to explore the use of unrelated volunteer donors. Initial results have demonstrated the feasibility of this approach. As a result, a national effort has begun to recruit HLA-typed volunteers in order to establish a registry of individuals who would be willing to serve as bone marrow donors. This manuscript explores the potential impact of establishing such a registry. We find that a registry of attainable size could more than double the number of marrow transplants now being performed. However, even with a registry of enormous size, it will still not be possible to identify an HLA-matched donor for some patients.

Bone Marrow Transplantation↗

PubMed1985

Marrow transplantation from related donors other than HLA-identical siblings.

Marrow transplantation has generally been limited to patients with a sibling who is genotypically identical for HLA. In a study of the acceptable limits of HLA incompatibility, 105 consecutive patients with hematologic cancers who received marrow grafts from haploidentical donors (study group) were compared with 728 similar patients concurrently receiving grafts from HLA genotypically identical siblings (control group). The unshared haplotypes differed variably: 12 were phenotypically but not genotypically identical for HLA-A, HLA-B, and HLA-D; 63 differed at one locus (A, B, or D); 24 at two loci; and 6 at three. A higher proportion of study patients had delayed engraftment, granulocytopenia, or graft rejection. Acute graft versus host disease occurred earlier and with greater frequency in study patients. The risk of the disease did not correlate with disparity for Class I (A or B) versus Class II (D-region) loci. Thus, incompatibility for HLA has an important effect on the course after clinical marrow transplantation. In spite of these complications, there was no statistically significant difference in the survival of the study patients and control patients who received their transplants during remission.

Acute Disease↗

PubMed1985

Electrophoretic variation between class II molecules expressed on HLA-DRw8 homozygous typing cells reveals multiple distinct haplotypes.

Two-dimensional (2D) gel electrophoresis of immunoprecipitated HLA-DR antigens from eight homozygous typing cells (HTC) expressing the HLA-DRw8 specificity revealed a clustering of polymorphic beta chain patterns into distinct electrophoretic variants. The variant patterns correlate with three discrete HLA-D clusters that are defined in the mixed leukocyte culture reaction (MLR) using DRw8-positive HTC. These HLA-D clusters have been provisionally designated Dw"8.1", detected primarily in Caucasoids, Dw"8.2", detected primarily in American Indians, and Dw"8.3", detected predominantly in Orientals. All three HLA-Dw"8.1" cell lines express a single DR-locus product as defined by immunoprecipitation with a DR-specific monoclonal antibody, P4.1. This DR beta chain is identical among the Dw"8.1" cell lines and different from the DR beta chains of the Dw"8.2" and Dw"8.3" cell lines. Two separate Dw"8.2" HTC express a shared DR beta chain that is slightly more basic than the 8.1 DR molecule; interestingly, one of these lines also expresses an additional DR-like beta chain not found in the other cells. Thus, the two lines defining the Dw"8.2" cluster share one distinct class II molecule, but differ in another and therefore are not biochemically HLA-identical. Cells from the Dw"8.3" cluster are likewise distinct from all other Dw8 clusters. One additional DRw8-positive HTC has been analyzed and found to be distinct from the Dw"8.1", "8.2" and "8.3" clusters by both MLR and 2D gels. Immunoprecipitates using monoclonal antibody 1B5 [anti-DR and anti-DQ(DS)] identify additional polymorphic class II variants among the cell lines tested. These data indicate that HLA-DRw8 is a public serologic specificity present on class II molecules expressed on multiple distinct haplotypes. These haplotypes differ from each other in expression of polymorphic class II molecules encoded by at least two HLA loci. They also differ in HLA-D, even though they all type as HLA-DRw8 homozygous. In Dw"8.2", variation in expressed beta chains is not reflected in variation in HLA-D, indicating that MLR, as well as serologic typing, does not detect the full degree of allelic polymorphism within HLA.

Antibodies, Monoclonal↗

PubMed1985

Definition of DW8.2 by primary and secondary mixed lymphocyte cultures.

Using HLA-DW8 homozygous typing cells (HTC) of different ethnic origin it is possible to identify three subgroups of the DW8/DRW8 product (Mickelson et al., 1983). To further characterize the DW8.2 subgroup defined by HTCs of Amerindian origin we have now generated bulk PLTs within members of one extended Amerindian family and within selected HTCs of Caucasian, Oriental, and Amerindian origin. A panel of 61 DRW8 positive and negative donors of the three ethnic groups was used to test 15 different PLTs. Our results demonstrate that it is possible to generate DW8.1, 8.2, or 8.3 sensitized lymphocytes which distinguish in secondary cultures between each of the three subgroups of the DW8/DRW8 products. Of 40 DRW8 cells tested, 100% Caucasians typed as DW8.1, 100% Amerindians were 8.2; 75% Orientals were DW8.3; 8.3% were DW8.2, and 16.6% could not be classified within any of these subgroups. DRW8 individuals of mixed ethnic origin typed as either DW8.1 or DW8.2 and one DRW8 homozygous donor behaved as heterozygous 8.1/8.2. These results confirm the subdivision of the DW8/DRW8 product and explain the poor correlation and unexpected responses reported in MLC with DW8 HTCs and DRW8 donors of different ethnic origin.

Asian People↗

PubMed1984

Immune responses in an untransfused patient with aplastic anemia: analysis of cytolytic and proliferative T cell clones.

Unexpected unidirectional reactivity was noted in MLC by the cells of an untransfused patient with aplastic anemia against cells from her genotypically HLA identical brother. To analyze this reactivity, lymphocytes from the patient were primed in vitro for six days with irradiated lymphocytes from the HLA identical brother and then cloned by limiting dilution in the presence of interleukin-2. Following a period of clonal expansion, the patient's cells were tested for specific proliferative (PLT) and cytolytic (CTL) activity against cells of the brother. Thirty-six clones demonstrated proliferative activity, 30 clones demonstrated cytolytic activity, and 114 clones showed neither. No clone demonstrated both cytotoxic and proliferative activity. Several patterns of specificity were seen for the cytolytic T cell (CTL) clones, including both allo- and autoreactivity. Two distinct patterns of specificity were noted for the proliferative clones: one reactive to cells from DR3-positive males; the other reactive only to cells from certain DR2-positive males and females. The DR3-restricted clones are presumably directed towards the H-Y minor histocompatibility antigen while the DR2-restricted clones are directed toward an undefined minor histocompatibility antigen. It is thus possible to isolate both alloreactive and autoreactive T cells from the peripheral blood of some untransfused patients with aplastic anemia.

Anemia, Aplastic↗