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Biomedical subjects

N L Reinsmoen

Publications and source records attributed to N L Reinsmoen.

At least 19 recordsLinked to original sources
PubMed1992

Differentiation of class I- and class II-directed donor-specific alloreactivity in bronchoalveolar lavage lymphocytes from lung transplant recipients.

Previous studies have demonstrated that donor antigen-specific primed-lymphocyte-test (PLT) reactivity of bronchoalveolar lavage lymphocytes is strongly associated with acute pulmonary rejection and with obliterative bronchiolitis (OB); however, a systematic analysis of PLT reactivity as being class I-or II-directed has not been performed. To assess reactivity directed against individual class I or II antigens, we tested a total of 67 BAL-derived lymphocyte samples from 26 recipients for alloreactivity in the PLT, using a pool of allogeneic cells and selected homozygous typing cells (HTCs) representing the HLA class I and II antigens expressed by the recipient and donor cells. The results obtained by PLT were correlated with the clinical status of the recipient with regard to rejection, infection, and OB. In 9 of 10 cases where transbronchial biopsy results were consistent with rejection, donor antigen-specific allogeneic PLT reactivity was observed and, more specifically, could be determined to be directed toward donor class II antigen in 8 of these cases. For 3 of 4 recipients tested chronologically, positive donor antigen-specific PLT reactivity was observed at the time of and 2-3 1/2 months prior to the diagnosis of rejection by transbronchial biopsy. During periods of acute infection, donor antigen-specific PLT reactivity was not observed; instead, non-specific PLT reactivity of BAL-derived cells (i.e., reactivity that did not correlate with any defined HLA antigens) was observed as well as reactivity associated with the self-antigens expressed by the recipients' cells. The PLT reactivity of BAL-derived cells from a recipient diagnosed with OB correlated specifically with one of the disparate donor class I antigens (HLA-B44). In 23 cases, BAL cells were propagated in the presence of autologous cells and rIL-2, thereby allowing for sufficient numbers of cells to test with a panel of 29 HTCs and to analyze for cell surface phenotype. The cultured BAL cells from 4 recipients undergoing a rejection episode demonstrated a predominant CD4+ phenotype consistent with the class II-directed reactivity observed in PLT. However, these results did not demonstrate a phenotype distinctive from the 7 BAL results obtained from 4 quiescent recipients. In marked contrast, the cultured BAL cells obtained from 4 recipients diagnosed with OB demonstrated a predominant CD8+ phenotype, with 60-92% of the cultured cells being CD8+. These results are consistent with the class I-directed reactivity observed in PLT.(ABSTRACT TRUNCATED AT 400 WORDS)

Adolescent↗
PubMed1991

Heterogeneity of the T-cell receptor beta gene rearrangements generated in myelin basic protein-specific T-cell clones isolated from a patient with multiple sclerosis.

Seventeen T-cell clones derived from the peripheral blood of a patient with multiple sclerosis and reactive with a synthetic peptide corresponding to residues 152-170 of the human myelin basic protein molecule were previously shown to be cytotoxic for myelin basic protein-coated target cells. Genetic restriction studies have now demonstrated that these clones recognize myelin basic protein in association with human leukocyte antigen DRw13. Studies of the T-cell receptor beta gene rearrangements generated by these clones demonstrated 12 different patterns, as evaluated by Southern blot analysis. Thus, the human T-cell response to myelin basic protein is exceedingly heterogeneous, even among T cells that recognize the same small fragment of the molecule in association with the same class II restriction element.

Clone Cells↗
PubMed1990

Functional analysis of MHC class II-restricted T cells derived from a Caucasian with a DR4, Dw15, DQw8 haplotype.

Rabies virus-specific CD4+ T lymphocyte clones were isolated from a Caucasian male vaccine recipient (DR4/7, DQw2/w8; DPw4) and studied for their major histocompatibility complex restricting elements. None of the rabies-specific T-cell clones could be induced to proliferate to antigen by either lymphoblastoid cells or DR-transfected L cells expressing DR4 molecules of the Dw subtypes commonly found on Caucasian individuals (Dw4, Dw10, Dw13, Dw14). The HLA-Dw subtype of the rabies vaccine recipient was determined by conventional mixed lymphocyte culture, and the results revealed that this individual had a DR4 (Dw15), DR7 (Dw7) phenotype. The presence of the DR4, Dw15 antigen was confirmed by nucleotide sequencing of the DR4B1 gene corresponding to the DRB1*0405 allele. Significant antigen-induced T-cell proliferative responses were obtained with two DR4, Dw15, DQw4 homozygous lymphoblastoid cell lines of Japanese origin (HAS-15 and KT-3) and with a L-cell transfectant expressing the DR4, Dw15 molecule. The existence of the DR4, Dw15 antigen in the Japanese has been reported to be associated with the DQw4 specificity. However, the presence of DQw8 (previously designated DQw3.2) and the absence of DQw4 in the lymphoblastoid cells of the Caucasian rabies vaccine was confirmed with monoclonal antibodies IVD12 (anti-DQw7 + DQw8 + DQw9) and HU46 (anti-DQw4) and by the reactivity of a DQw8-restricted antigen-specific T-cell clone. These studies indicate, contrary to previous findings, that the DR4, Dw15 molecule may be present in Caucasian (non-Japanese) individuals in association with DQw8.

Alleles↗
PubMed1990

Structural model for T-cell recognition of HLA class II-associated alloepitopes.

In an effort to investigate the structure-function relationship of HLA class II molecules vis-à-vis alloepitope expression, cloned T-cell reagents were used to define polymorphic epitopes associated with DR and DQ molecules. DNA sequences of genes encoding allelic or isotypic DR or DQ molecules that appear to express the same T-cell-defined epitopes were compared in an attempt to identify association of shared sequences with shared epitopes. When sequence sharing is associated with shared epitope expression, we suggest that it is the shared sequence that encodes the epitope in question. Based on the hypothetical three-dimensional structure of the class II molecule, an approximation is made as to which parts of the HLA class II molecule are involved in alloepitope expression. T-cell clones were generated from cells primed against HLA-DR2 haplotypes representing the cellularly defined subgroups Dw2 or Dw21 (previously designated MN2, FJ0, or Tb24). Those clones determined to be DR- or DQ-directed based on monoclonal antibody inhibition assays were tested by panel cell analysis utilizing DR2-positive and DR2-negative target cells. The data support the concept that amino acids 67, 70, 71, and 74 for DR molecules and amino acids 57, 70, and 71 for DQ molecules, which appear to comprise one face of the alpha helix, are of primary importance in T-cell recognition. In other cases, sharing of both the second hypervariable region (amino acids 25-33) and the third hypervariable region (amino acids 67-74) appears necessary to explain epitope sharing for DR molecules. We emphasize that the involvement of these two hypervariable regions may indicate that alloepitope expression involves the complex of class II molecule plus peptide, with the second HVR primarily involved in determining which peptides are bound and the third in T-cell receptor (TcR) recognition and/or peptide binding; we do not rule out that conformational changes of the second HVR can induce conformational changes in the third HVR. Finally, shared alloepitopes detected by some clones could not be explained based on shared primary sequences.

Amino Acid Sequence↗
PubMed1990

The impact of HLA matching on graft survival and on sensitization after a failed transplant--evidence that failure of poorly matched renal transplants does not result in increased sensitization.

There are costs (both financial and ethical) to distributing kidneys by HLA-match (time, transportation, repeat crossmatch; possibly bypassing a more deserving recipient). Arguments favoring matching include better short- and long-term survival, and decreased panel-reactive antibody (PRA) if a well-matched vs. poorly matched transplant fails. We studied these phenomena in a single institution. Since 1970, 1329 patients received cadaver (CAD) transplants; for those with defined antigens (n = 1316) there was no difference in 10-year graft survival in those with a less than or equal to 1 AB match vs. those with greater than 1 AB match or those with less than or equal to 1 AB mismatch (mm) vs. greater than 1 AB mm. Similarly there was no difference in those with less than or equal to 2 BDR mm vs. greater than 2 BDR mm. In fact, those with less than ABDR mm had worse 10-year graft survival (55%) than those with greater than or equal to 3 ABDR mm (61%) (P = .001). For patients with function greater than 6 months there was no difference in long-term outcome based on HLA match or mm. These findings were similar for patients both with and without CsA immunosuppression, and for primary and retransplants. A total of 382 patients transplanted since 1980 have lost their grafts (146 died with function). All had received pretransplant transfusions. Of 236 alive after graft loss, 64 had no postgraft failure PRAs (22 out of state, 23 chose to remain on dialysis, 19 died less than 3 months after graft loss); 172 had PRAs after failure; 106 (62%) have been retransplanted. Mean peak PRA in those retransplanted was 23 +/- 31 (range 0-100) vs. 46 +/- 39 (range 0-100) in those not retransplanted (P less than .05). Patients were stratified by PRA prior to first transplant (0%, 1-20%, greater than 20%). For recipients with 0% PRA, failure of a CAD transplant (n = 58) was no more likely to result in an increase of PRA than failure of a living-related donor (LRD) transplant (n = 49) (NS). For those with an increase, mean increase was 45% +/- 34 after LRD transplant and 41% +/- 28 after CAD transplant (NS). The proportion developing PRA greater than or equal to 60% was not different after a failed LRD (7/49) or CAD (11/58) transplant (NS). Other subgroups had similar results. AZA or CsA immunosuppression did not affect development of increased PRA after a failed graft.(ABSTRACT TRUNCATED AT 400 WORDS)

Antibody Formation↗
PubMed1990

A new in vitro approach to determine acquired tolerance in long-term kidney allograft recipients.

Previous studies indicate some kidney allograft recipients treated with total lymphoid irradiation, cyclosporine, or conventional immunosuppressive therapy demonstrate specific proliferative unresponsiveness in mixed lymphocyte culture (MLC) to donor cells at various times posttransplant. To investigate possible donor-specific hyporeactivity, we have studied 3 patients treated with TLI whose grafts have survived longer than 10 years; 2 patients given the same immunosuppressive protocol but without TLI whose grafts have survived longer than 10 years; and 27 CsA-treated living-related donor and cadaver-allograft recipients 1 year posttransplant. We confirmed previous observations of hyporeactivity of some patients' cells to stimulation by donor cells. In addition, we identified hyporeactivity to stimulation by homozygous typing cells (HTCs) defining the HLA-Dw specificities of the donor cells for all 3 of the 3 TLI patients, 1 of the 2 non-TLI patients, and 9 of the 27 patients 1 year posttransplant. The LRD recipients with donor-specific hyporeactivity as defined by the HTC analysis demonstrated fewer rejection episodes (25% vs. 57%) and lower mean creatinine levels (1.18 vs 1.78 mg/dL) than patients without donor-specific hyporeactivity. These studies demonstrate the feasibility of monitoring the immune status of allograft recipients posttransplant by means of HTC analysis, eliminating the need for pretransplant specimens. This approach provides a possible means to assess which patients may have acquired donor-specific hyporeactivity to their kidney allograft and thus may require less immunosuppression.

Clone Cells↗
PubMed1987

Determination of DR beta 1 alleles of DR4/Dw subtypes by oligonucleotide probing.

The extensive Dw polymorphism of HLA-D has, to date, been defined using cumbersome and lengthy techniques involving T-cell proliferative reactions. These techniques have not necessarily provided information about the genotype at any single locus of HLA-D because the products of more than a single locus of HLA-D (e.g., DR beta 1 and DR beta 2 of the DR2 haplotypes and in some cases DQ) can contribute to T-cell proliferation and thus to assignment of Dw phenotype. We have recently reported that DR beta 1 alleles relating to the Dw polymorphism can vary by only a single, or a very few, nucleotide differences; the products of this locus are recognized in allo- and restricted responses. Given the importance of defining this polymorphism, we have used specific oligonucleotides as hybridization probes to differentiate between these very closely related alleles. This approach provides the basis for a rapid method for HLA-D typing which, as additional sequence information becomes available, will likely be generally applicable. In addition, oligonucleotide probing allows definition of a gene transcript from an individual locus of HLA-D rather than the phenotype results provided by Dw typing.

Alleles↗
PubMed1987

T-cell clonal analysis of HLA-DR2 haplotypes.

Individuals carrying the serologically-defined DR2 haplotypes can be further subdivided by utilizing T cell response to define several HLA-Dw/LD clusters: Dw2, Dw12, LD-5a, and a related group composed of FJO, AZH, and MN2. We have used cytotoxic T cell clones generated against these DR2 haplotypes to define the DR and DQ associated Dw/LD polymorphisms. Clones were categorized as DR, DQ, DP, or class I-directed based on monoclonal antibody (MoAb) inhibition studies. We analyzed the distribution of the determinants detected by these clones based on analysis with panels of cells. Some DR-directed clones only lysed cells positive for the Dw subtype of the sensitizing cell. A few DR-directed clones lysed most DR2 positive cells. Other DR-directed clones detected determinants shared by cells of different subtypes: determinants shared by Dw2 and Dw12 positive cells and determinants shared by MN2, FJO, AZH, and LD-5a positive cells. The majority of the DQ-directed clones only lysed cells positive for the sensitizing HLA-Dw subtype. A group of clones primed against an LD-5a HTC, whose cytolytic activity was not blocked by any monoclonal antibodies tested, lysed some, but not all, DR2 positive cells as well as Dw1 and Dw10 positive cells. The results suggest the detection of determinants on these molecules which may be shared by cells of specific Dw subgroups and determinants shared by most cells of the sensitizing serological specificity. The basis of this observed polymorphism and influence on the allogeneic response are discussed.

Antibodies, Monoclonal↗
PubMed1987

Protein analysis of HLA-DR2.3 haplotype.

The human major histocompatibility antigens (HLA-DR, HLA-DQ, and HLA-DP), consisting of alpha and beta chains, show extensive polymorphism. Based on differences of T-cell responses, the serologically defined DR2 specificity has been divided into Dw2, Dw12, and several putative additional "Dw subtype" specificities (AZH, FJO, and MN2, which we shall designate as "DR2.3", and LD-5a). To investigate the relationships of these specificities, DR and DQ molecules were analyzed by two-dimensional gel electrophoresis (2D-PAGE). The 2D-PAGE patterns of the DR molecules are identical among DR2.3 cells. Three DR beta chain polypeptides are expressed on DR2.3 cells. The electrophoretic mobilities of two of these beta chains are different from those of beta chains from the Dw2 and Dw12 cells. The DR molecules of the LD-5a cells show similar 2D-PAGE patterns with those of DR2.3 cells. The 2D-PAGE patterns of DQ molecules are identical among DR2.3 cells. The electrophoretic mobilities of DQ beta chains are different for Dw2, Dw12, LD-5a, and DR2.3 cells. These results indicate that the AZH, FJO, and MN2 cells have identical or very similar DR and DQ molecules and constitute a third, and relatively homogeneous, subgroup ("DR2.3") of the DR2 specificity. In addition, DR2.3 cells have a unique characteristic in that they express three DR beta chains.

Antibodies, Monoclonal↗
PubMed1986

Clonal analysis of HLA-DR and -DQ associated determinants: their contribution to Dw specificities.

In order to investigate the distribution of epitopes recognized by T-cell clones directed against HLA class II products, bulk primed cell populations were generated using cells matched for class I determinants but disparate for class II determinants. Cells were cloned by single cell deposition (FACS IV) or limiting dilution (1 cell/3 wells), and assayed for proliferative and cytolytic function with panels of well-characterized cells. All cytolytic clones generated from an anti-DR4/Dw4/DQw3 priming combination or an anti-DR2/Dw2/DQw1 priming combination lysed essentially all targets sharing the same Dw type as the sensitizing cell. In some cases, other targets were also lysed. For instance, some clones were lytic to targets bearing the same DR antigen but another Dw subtype including a few clones lytic to virtually all cells carrying that DR specificity. An occasional target cell expressing a different DR antigen from the sensitizing cell was also lysed by these clones, in some cases to the same extent of lysis seen on the specific target. Monoclonal antibody inhibition studies identified three groups of clones: the DQ directed clones and clones apparently directed at more than one DR product. However, the number of molecules detected for each haplotype remains to be investigated. Our data indicate that determinants detected on both DR and DQ products are associated with the Dw type of the sensitizing cell showing that there is polymorphism recognized by T cells on both DR and DQ that is subtypic to the serologically defined specificities. Thus, it appears that the bulk T-cell response is a composite of individual clones recognizing distinct determinants on these class II molecules. The implications of these findings for studies of HLA restricted recognition are discussed.

Antibodies↗
PubMed1985

A new HLA-DP (SB) specificity (Cp63) defined in Japanese and Caucasian cell populations by cloned PLT cells.

Cp61, Cp62, and Cp63 are HLA-DP (SB) specificities detected by PLT testing using cloned PLT cells derived from Japanese cell donors. It was confirmed, using Japanese and Caucasian cell panels, that Cp61 is identical to DPw5 and Cp62 is identical to DPw4. Cp63, however, was found not to correlate with any known HLA-DP specificities. The gene frequency of Cp63 was found to be 3.6% in Japanese (N = 42) and 1.9% in Caucasian (N = 52) cell panels of limited size. No triplet assignment of DPw specificity including Cp63 was found in the Japanese and Caucasian cell populations. The relationship between the Cp63 and the HLA antigens demonstrated no significant correlation with HLA-A,B,C,DR, or MT antigens in this study. Linkage between Cp63 and HLA was confirmed from family segregation studies. This is the first report of a new DP specificity termed Cp63.

Asian People↗
PubMed1985

Analysis of human class II antigens by cloned cytolytic T cell reagents: a study using HLA loss mutant lymphoblastoid cell lines and monoclonal antibodies detecting the HLA-DP product(s).

We have utilized cloned T cell reagents and ionizing radiation-induced mutants of an HLA heterozygous lymphoblastoid cell line (LCL) to investigate the determinants detected by the cell-mediated lympholysis (CML) assay. Cells of an LCL clone, 721.501, an HLA haplotype loss mutant expressing the HLA-A2-Cw1-Bw51-DR1-Dw1-DQw1-DPw2-GLO haplotype were used as sensitizing cells for responder cells in vitro. "Cloned" reagents were generated by single-cell deposition of cells of a bulk reagent primed against 721.501 cells. Those clones were screened for cytolytic activity against HLA loss mutant targets (derived from LCL 721) of four different categories; HLA-A2 loss only, A2-Cw1-Bw51 loss, A2-Cw1-Bw51-DR1-DQw1 loss, and the entire HLA haplotype loss. Of 196 clones tested, 36 were cytolytic, including three anti-A2, five anti-Bw51/Cw1, 12 anti-DR1/DQw1, 13 anti-DP region associated with DPw2, and three of undetermined specificity, based on cytolytic patterns against the HLA loss mutant targets. Of 25 anti-HLA class II lytic clones, 23 (92%) fitted the characteristics of helper cell-independent cytolytic T cells (HITc), whereas only two of eight (25%) anti-class I clones were HITc. The 13 anti-DP region clones were divided into three subgroups defined by blocking by anti-FA and not Tü39 monoclonal antibodies (MoAb), by Tü39 and not anti-FA, and by both MoAbs.

Antibodies, Monoclonal↗
PubMed1979

Anomalous mixed lymphocyte culture reactivity between HLA--A, --B, --C, --DR identical siblings.

Complete HLA typing including HLA--A, --B, --C, --DR (D related B cell typing), --D, mixed lymphocyte culture (MLC), and primed lymphocyte testing (PLT), together with complete red blood cell (RBC), glyoxalase (GLO), GBG (Factor B), and phosphoglucomutase 3 (PGM3) typings were performed on a informative family. The five siblings inherited the four possible combinations of parental HLA haplotypes, and two of the siblings were HLA--A, --B, --C and --DR identical. Repeated MLC testing of the family revealed positive mixed lymphocyte reactivity in all combinations. B cell typing for the DR specificities demonstrated no variation from the expected inheritance pattern and specifically no recombination event. GBG and GLO typings militated against a recombination involving the paternal chromosome. HLA--D testing revealed that only one of the HLA--A, --B, --C, and --DR identical siblings gave typing responses to the HLA--Dw3 specificity present on that maternal haplotype. Utilizing HLA haploidentical combinations, lymphocytes were primed against the four parental haplotypes and the non-Dw3 haplotype of interest (Aw24--B8--DRw3--LDY) in the PLT. The sibling inheriting this haplo-type did not restimulate cells primed against the A2--B40--DRW6--LDY specificity. Furthermore, no discrimination was observed in the restimulation of lymphocytes primed against this haplo-type. Possible interpretations of these family data include: a spontaneous mutation, non-major histocompatibility locus (MHC) stimulation, and HLA--DR/D recombination.

B-Lymphocytes↗
PubMed1978

Antigens associated with acute leukemia detected in the primed lymphocyte test.

Mixed lymphocyte culture (MLC) studies of families with several leukemia patients, all potential bone marrow transplant recipients, demonstrated that cells from acute myelogenous leukemia patients (5 of 5) and acute undifferentiated leukemia patients (1 of 4) in relapse stimulated autologous lymphocytes as well as lymphocytes from siblings known to be identical at the major histocompatibility linkage group. In the patients studied, the blast transformation induced by leukemia cells was not detectable when the patient was in remission. Stimulation by leukemia cells also elicited increased responses of the lymphocytes from normal haploidentical siblings, parents, and unrelated individuals as compared to stimulation by normal allogeneic cells or leukemia cells of patients with leukemia in remission. The primed lymphocyte test (PLT) was used successfully to establish HLA-D identity of the leukemia patients and their respective HLA-identical siblings, despite high percentages of circulatory blasts. Utilizing lymphocytes from normal siblings primed against the leukocytes from an HLA-identical sibling with leukemia, we also presented results of PLT's which suggested that the stimulation induced by leukemia cells in MLC was produced by leukemia-associated antigens.

Acute Disease↗