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Biomedical subjects

W Leibold

Publications and source records attributed to W Leibold.

At least 19 recordsLinked to original sources

Immunological characterization of mononuclear cells in peripheral blood and regional lymph nodes of breast cancer patients.

Mononuclear cells from peripheral blood and draining lymph nodes of 40 patients with invasive locoregional breast cancer were examined for immunological cell surface markers (E, EAhuman, EAox, EAC, SIg pos.). Concomitantly, blood lymphocytes from 36 healthy women and axillary and mesenteric lymph-nodes from patients without malignant diseases were tested as controls. In peripheral blood of tumor patients E rosette-forming cells were slightly diminished as compared to the control group, whereas EAox and EAC rosette-forming cells were increased. These differences may be age-dependent rather than tumor-related. In the draining lymph nodes of breast cancer patients as well as in the control lymph nodes, the percentages of EAC rosette-forming cells and SIg positive lymphocytes were significantly increased compared to peripheral blood, whereas E and EAhuman rosette values remained unchanged. Percentages of EAox rosettes on the other hand were strongly diminished in the draining lymph nodes, suggesting that the EAhuman and EAox rosetting techniques detect 2 types of Fc-receptor bearing cells. No significant differences were found between the cell surface marker analysis of tumor-free and metastatic lymph nodes of breast cancer patients and the control lymph nodes.

Adult

HLA-D locus typing in ankylosing spondylitis and Reiter's syndrome.

HLA-D typing of 44 patients with ankylosing spondylitis (AS) and 31 patients with Reiter's syndrome (RS) did not show increased frequency of any particular Dw allele in either population of patients as compared to controls. Such studies also allowed each patient's general response to be compared with other general responses within each experiment. Contrary to reports of diminished lymphocyte responses in AS patients, hyperresponsiveness in both AS and RS patients was found.

Alleles

HLA-D typing with lymphoblastoid cell lines. VI. Rationale and goals of data reduction.

This report documents various characteristics of HLA-D typing by mixed leukocyte culture reactions when lymphoblastoid cell lines (LCLs) are substituted for peripheral blood lymphocytes as the stimulator cells. It also provides the rationale for designing the computer program described in the subsequent report. In such experiments, each donor to be HLA-D typed is stimulated with a panel of 30-50 HLA-D homozygous LCLs, each defined HLA-D allele being represented by several different human homogygous typing cells (LCT-HTCs). Variability in the strength of eahc donor's general response and in the strength of stimulation by each LCL-HTC makes it necessary to normalize raw data before the responses of various combinations can be compared and typing responses distinguished from non-typing responses. The autologous response and its equivalent effect among allogenetic combinations, the so-called "autologous-stimulation" effect, must also be distinguished from true allogeneic responses. The latter has been accomplished by "modelling," as described in the subsequent report. EBV-negative donors can also be HLA-D typed by this method despite the EBV-positivity of the LCL-HTCs. Preliminary analyses suggest that the HLA-D alleles defined by this method appear to segregate with appropriate haplotypes in family studies.

Alleles

HLA-D typing with lymphoblastoid cell lines. VII. A computer program for data analysis.

When lymphoblastoid cell lines (LCL) are substituted for peripheral blood lymphocytes from human typing cell donors in HLA-D typing experiments, a data analysis program must be designed to distinguish the effect of allo-reactivity from those peculiar to LCL, mainly the "autologous-stimulation" effect. The computer program described in this report was created specifically for such an analysis. The rationale for the design of this program is presented in the preceding report (see this issue).

Cell Line

Suppression of lymphoblastoid cell line proliferation by antisera to HLA-DR and other HLA antigens.

We have observed that certain antisera to HLA antigens suppress the in vitro proliferation of lymphoblastoid cell lines. An antiserum to HLA-B8 demonstrated suppressor activity associated with the presence of B8 antigen on the target LCLs; this activity was removed by absorption with platelets or with B8-positive LCLs. An antiserum to HLA-DRw7 suppressed proliferation of all six DRw7-positive LCLs and none of 12 DRw7-negative LCLs; this activity was removed by absorptions with three DRw7-positive LCLs, each representing a different HLA-D allele (i.e. Dw7, 10 and 11); it could not be removed by triple serial absorptions with platelets from multiple-donor pools. These experiments indicate that the suppressor activity of this serum is specifically associated with antibodies to DRw7. Our model of LCL growth suppression by HLA antisera is easily manipulated and provides a definitive tool for further characterizing HLA antigens and antisera in a complement-independent system.

B-Lymphocytes

HLA--D typing with lymphoblastoid cell lines. IV. Allelic relationships.

Responses of approximately 200 persons were measured by mixed leukocyte cultures against 41 lymphoblastoid cell lines which were thought to be homozygous for HLA--D (i.e. LCL-HTCs). These responses were standardized by computer analysis and the resultant Interaction Indices for each LCL-HTC were compared with those of every other LCL-HTC in a correlation matrix of 861 independent analyses. Only those analyses involving LCL-HTCs of defined HLA--D types are reported here. In general, these studies confirmed many allelic relationships of reputedly similar LCL-HTCs; however, several unexpected groups of correlations suggested that groups Dw1 and 3 overlap to form Groups "1a-3," "1b-3" and "true 3." The "1b-3" group also included Dw8. Segregation of the "1b-3" allelic group was also observed in a family study. These preliminary analyses suggest a working model for further investigation of HLA--D allelic relationships (Fig. 3). On the other hand, they may also be interpreted to suggest that "typing" with LCL-HTCs may be controlled by a separate HLA locus which is closely related to, but distinct from, HLA--D.

Alleles

75Se-release: a short and long term assay system for cellular cytoxicity.

The gamma-emitting aminoacid 75Se-selenomethionine (75SeM) was examined as a target cell label in cytotoxic assays. It was efficiently taken up by activated, intensively metabolizing cells of various types but hardly at all by resting or low-metabolizing cells. Culturing activated cells in methionine-deficient medium with 3--5 mu Ci75SeM/ml for 18--22 h usually resulted in an uptake of 3--20 cpm/cell which was 3--200 times that of 51Cr marked cells. 75SeM-labelled cells kept in medium at ambient temperature or at 37 degrees C, maintained a high radioactivity per cell and a viability above 85% for at least 72 h without significant increase in spontaneous isotope release or loss in sensitivity in subsequent cytotoxic tests. 75Se-labelled material released from target cells was not reutilized by unlabelled lymphoid cells. Provided the cells were carefully washed after labelling and kept in optimal culture conditions, the reasonably low baseline release (usually 0.6--1.8% of input/h) in the medium control allowed performance of long-term assays of up to 54 h. However, strong cytotoxic reactions (e.g. ADCC) could cause over 50% specific 75Se-release within 5 h. With constant amounts of effector cells (3.6 x 10(3) up to 3 x 10(5)/well) identical or even higher, specific releases were obtained on 6 x 10(2) targets as compared to 1 x 10(4) targets/well. Thus, the 75Se-release assay offers a single monitoring system suitable for short (3--6 h) and long term (usually up to 44 h) cytotoxic reactions on a microscale, using 1 x 10(3) or less targets/well. Its sensitivity permits evaluation of strong and weak reactions as well as early and delayed onset cytotoxicity. In addition, with a gamma-spectrometer the radioactivity of 75Se can easily be distinguished from that of 51Cr. Due to this, and an improved method for 51Cr labelling of cells (10 mu Ci 51Cr/ml medium for 18--22 h), a double gamma-labelling of cellular proteins is available which provides new possibilities for monitoring cellular interactions in short and long term tests.

Animals

New groups and segregant series among B-cell alloantigens of the Merrit system. A study of leukemia cells, peripheral B cells, and lymphoblastoid cell lines.

On the basis of reactions with a chronic lymphatic leukemia cell panel, evidence for 6 new specificities of the Merrit B-cell alloantigenic system in man is presented, bringing the number of provisionally defined specificities to 19. These can be roughly divided into two segregant series. The system is well represented on both homozygous and heterozygous B-type lymphoblastoid cell lines. In the cell lines some specificities show a suggestive but inexact correlation with HLA-D locus factors. This correlation is represented also on peripheral blood B cells, on the non-T variety of acute lymphatic leukemia cells, and on acute myeloid leukemia cells. Although great similarities exist, each of the cell populations may manifest differences other than or in addition to mere differences in B-cell antigen frequencies.

B-Lymphocytes

Reactivity of alloantibodies of the Merrit B-cell system with leukemic cells and lymphoblastoid cell lines.

Twenty lymphoblastoid cell lines were typed with 38 antisera defining the 13 groups of the Merrit B-cell alloantigenic system, and results were compared with those of a 20-member CLL cell panel. While overall pattern of Merrit groups for the two types of cell population was similar, some sera in a number of groups showed unexpected negative cytotoxic reactions with lymphoid cells compared to other sera of the same group. However, in a number of instances, these negatively reacting cells could still specificity absorb out tb Merrit antibodies, suggesting that lymphoblastoid lines may show a higher incidence of the CYNAP (cytotoxicity-negative-absorption-positive) phenomenon than CLL cells. Twelve of the lymphoblastoid lines were HLA-D homozygous and in addition, each cell line displayed positive reactions for no more than 2 of the 13 Merrit groups, while 4 of the 8 heterozygous lines were positive for 3 of the 13 groups. These findings indicate that HLA-D-homozygous cell lines are antigenically simpler than the heterozygous lines for the Merrit system and suggest that HLA-D-homozygosity may correlate with homozygosity for at least one of the Merrit loci.

B-Lymphocytes

Simplification of B-cell antisera of the Merrit system with platelets and lymphoblastoid cell lines.

Human sera with anti-HLA and anti-Merrit B-cell reactivities were absorbed with different volumes of a pool of packed outdated human platelets obtained from 237 donors. After absorption, each serum was tested against a panel of CLL cells and normal unfractionated peripheral blood lymphocytes that had given positive reactions with the same serum prior to absorption. The optimal platelet volume for complete absorption of anti-HLA and full preservation of anti-Merrit B-cell activities in these sera was found to be 1/10 to 1/5 of the volume of serum. Higher volumes of packed platelets, especially over 1/1, in most instances reduced anti-B-cell activity "nonspecifically." Among the platelet absorbed sera, those showing no HLA and strong anti-Merrit B-cell cytotoxicity were used for cross-absorption studies with cultured human lymphoid cell lines. Multi-specific anti-Merrit sera could be simplified by this method. Cultured lymphoblastoid cell lines represent a valuable resource, since large volumes of cells may be needed for absorption and simplification of human sera containing allogeneic B-lymphocyte antibodies.

B-Lymphocytes