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D J Schendel

Publications and source records attributed to D J Schendel.

At least 73 records · Page 4Linked to original sources

Rare variant of complement C4 is seen in high frequency in patients with primary glomerulonephritis.

59 unselected patients with primary glomerulonephritis were phenotyped for alleles of the MHC-linked complement genes, C4A, C4B, and BF. A rare variant of the C4B locus, C4B*2.9, was found in 25% of these patients compared with only 2% of the normal population--a relative risk of 22.1 for glomerulonephritis in individuals with this variant. Subdivision of patients by histological classification of glomerulonephritis revealed a significant association of C4B*2.9 with the membranoproliferative form. There were no significant associations between primary glomerulonephritis or its subtypes and the other HLA markers tested.

Alleles↗

Clonal expression of differentiation and Ia-like antigens on alloreactive human T lymphocytes.

A series of human T lymphocyte clones showing specific responses for major histocompatibility complex-encoded alloantigens were isolated from a single culture. They were classified into distinct functional groups based on measurement of alloantigen-specific proliferation and cytotoxicity. Surface antigen expression on these clones was analyzed using a panel of monoclonal antibodies specific for T cell differentiation antigens and for Ia-like antigens. Four distinct groups were identified based on the Leu series of differentiation antigens: (a) Leu1+,2+,3+,4+; (b) Leu1+,3+,4+; (c) Leu1+,2+,4+; and (d) Leu2+,4+. Functionally distinct clones showed not only differential expression of Leu2 and Leu3 , antigens which had previously been shown to distinguish helper/inducer and cytotoxic/suppressor cells, but also of Leu1, an antigen classified as a common T cell marker. All of the clones were found to express Ia antigens as detected by three framework monoclonal antibodies, but none were found to express a determinant associated with the DR3 allospecificity , although it was found on normal B lymphocytes and a B cell line established from the HLA-DR3 individual from whom these clones were derived.

Antibodies, Monoclonal↗

Cultured human T lymphocyte lines and clones as immunogenetic tools.

Human T lymphocytes were sensitized in vitro to allogeneic determinants encoded by the HLA region and subsequently cloned by limiting dilution in the presence of human T cell growth factor. An in vitro priming protocol entailing multiple rounds of allostimulation in mixed lymphocyte culture, using a disparate stimulating to responding cell ratio (10:1), enabled a high frequency of functionally active clones to be isolated. The specificity of the proliferative and cytotoxic reactivities was ascertained directly by screening in an HLA characterized family.

Alleles↗

C4 allotyping distinguishes HLA-B14.1 and B14.2 subgroups.

Complement allotyping (C4, C2, and BF) was performed in 60 unrelated individuals and 15 families characterized for the subtypes (14.1 and 14.2) within HLA-B14. Eighty-seven percent of B14.2 individuals typed positive for the rare C4A2 variant. In contrast, less than 7% of B14.1 individuals were positive for this C4 allotype which is in keeping with a control background frequency. Family studies revealed that three distinct complotypes (complement haplotypes) are characteristic for the two HLA-B14 subgroups. The SC22 and FC31 complotypes characterize the B14.2 subtype, whereas SC31 appears to define B14.1.

Complement C4↗

MHC-linked class III genes. Analysis of C4 gene frequencies, complotypes and associations with distinct HLA haplotypes in German Caucasians.

The class III complement components, C4, C2 and factor B (BF), are encoded in the human major histocompatibility complex (MHC). The two genes determining C4 (C4A and C4B) display considerable polymorphism and, thus, are important markers for HLA. In combination with alleles of C2 and BF they can be grouped into unique complotypes. We have analyzed the C4 alleles in a panel of 204 unrelated German Caucasians and studied their segregation with HLA haplotypes in 24 normal families. Inclusion of the class III markers with the class I and II alleles provides a more refined picture of the genetic structure of the MHC in these families. When charted according to the HLA-B locus specificities the MHCs can be clustered into groups showing distinctly homogenous or heterogenous complotypes. The identification of such groups is valuable for the selection of genetic material to analyze the molecular genetics of the human MHC.

Complement C2↗

Biochemical and cellular identification of HLA-Bw44 subtypes.

Biochemical analysis of HLA-Bw44 antigens by two-dimensional gelelectrophoresis and one-dimensional isoelectric focusing (IEF) from serologically heterozygous and homozygous donors allows the identification of two distinct types of HLA-Bw44 molecules, designated as Bw44-I and Bw44-II. These results are in concordance with the data obtained by CML-typing where at least two types of Bw44 target cells can be distinguished clearly as well. The antigen of type I has a more acidic isoelectric point (IEP) than that of type II. The difference in IEP are not due to differences in sialic acid content.

Female↗

Direct demonstration of an HLA-DR allotypic determinant on the low molecular weight (beta) subunit using a mouse monoclonal antibody specific for DR3.

A murine monoclonal antibody directed against a human B cell surface antigen with the characteristics of HLA-DR is described. The antigen detected is tightly linked to HLA and is correlated with the alloantigen HLA-Dw/DR3. Reactivity with a fraction of Dw/DRw6 cells is also observed. The determinant recognized by this antibody has been shown to be present on the smaller molecular weight beta subunit of the HLA-DR antigen.

Animals↗

Production of human T cell growth factor.

We provide here a protocol for production of T cell growth factor (TCGF) using cells isolated from defibrinated blood. Whether combined in allogeneic pools or tested as single donors, these cells consistently yield high activity TCGF, following PHA stimulation. Protocols using cells isolated from heparinized blood have often included addition of indomethacin or removal of adherent cells, or both, to overcome the problem of frequent nonproducing cultures. Our failure to find nonproducing cultures using this source of cells suggests that inhibitory cells or factors are removed by the blood clot formed during defibrination. A distinct economic advantage is gained by using these cells, not only because of the reliability of obtaining active supernatants, but also because the same blood can be used for preparation of a serum pool for cell cultures and the erythrocytes are useful for absorption of contaminating PHA present in the TCGF-containing media. Not only can defibrinated blood leukocytes be stimulated by PHA to release TCGF, but when cultured in allogeneic mixtures containing no PHA, they also release active TCGF.

Culture Media↗

Noncodominant expression of target antigens recognized by human cytotoxic T lymphocytes.

Human CTL that recognized MHC-controlled determinants distinct from HLA-A. B, C, and D/DR antigens were tested in a family with nine siblings. Segregation analysis of positive CML reactions showed strong lysis of target cells of three HLA-identical siblings, inheriting the b and c MHC haplotypes (blc), but not of the parents or siblings inheriting only one of these haplotypes. Some cytotoxicity was seen against parental target cells, although it seemed to be qualitatively distinct from that directed against the b/c targets. Studies using the competitive inhibition technique showed that cells inheriting only the b or c haplotypes were not effective in decreasing the specific cytotoxicity. Furthermore, simultaneous inclusion of inhibitor cells of both the b and c haplotypes did not hinder the specific cytotoxicity. Induction of CTL recognizing this new determinant did not occur when either antigens of the b or c haplotypes were used as MLC stimulating cells, or when antigens of both haplotypes were presented simultaneously, but on separate stimulating cells, in a three-cell MLC. These results suggest that the target determinant recognized by these unusual CTL is complex; it may be formed through interactions of two surface molecules or by genetic complementation yielding a hybrid antigen.

Binding, Competitive↗

Isolation of HLA-haplotype-specific human T-lymphocyte clones.

Human T-lymphocyte clones showing proliferative and cytotoxic responses specific for HLA antigens were isolated. To obtain enriched numbers of functionally active colonies, the cells were first sensitized in vitro in a tertiary mixed lymphocyte culture (MLC), using 10 stimulating cells for each responding cell. To assess quickly the genetic specificity of the clones, the cells used in the priming MLC and later as screening cells were prepared from an HLA-typed family. Using this approach, 75% of the colonies seeded at 1 cell/well showed functional activity specific for HLA.

Cell Separation↗

Cell-growth kinetics and karyotype analysis of human memory lymphocytes responding to alloantigen and mitogen.

Peripheral-blood lymphocytes were primed in vitro with the mitogen phytohemagglutinin (PHA) or with allogeneic cells and their memory responses studied following sequential restimulation with either mitogen or alloantigen. Chromosome preparations were made every 24 hours following exposure to the stimulating agents. Cultures were labeled with BUdR for sister-chromatid staining of the chromosomes which provided information about the kinetics of cell growth and rates of sister chromatid exchange. Cultures containing n BUdR were used for the investigation of cell karyotypes after chromosome-banding. Following PHA as well as alloantigen restimulation, an earlier reaction of the responding cells was observed. The peak response after the first stimulation was found at 120 h with allogeneic stimulation and at 60 h with mitogen stimulation. In the second round of stimulation, the peak occurred after 48 h (allogeneic) and 36 h (PHA) and following the third stimulation after 36 h (allogeneic) and 24 h (PHA). The speed of cell growth was decreased following restimulation with either alloantigen and mitogen. In contrast to the allogeneic restimulation, the number of cells responding after PHA restimulation was decreased. No systematic numerical or structural aberration of the karyotype was detected following repeated stimulation with either alloantigen or mitogen. In this sense, the lymphocyte subpopulations selected by repeated stimulation did not differ from the starting material. On the other hand, the sister-chromatid exchange (SCE) frequency was increased following allogeneic restimulation, whereas it remained constant with PHA restimulation.

Humans↗

Human T cell differentiation antigens characterizing a cytotoxic/suppressor T cell subset.

Monoclonal antibodies were raised against the leukemic T cells from a patient with chronic lymphocytic leukemia. Two antibodies, termed T411 and T811, were obtained which were reactive by indirect immunofluorescence only with cells of the T cell lineage. The T411 antibody recognized a polypeptide chain of 100,000 dalton apparent molecular weight which was present on the surface of 94 +/- 4% of peripheral blood T lymphocytes, but only on 20 +/- 8% of thymus cells. The antibody T811 reacted with a surface molecule composed of 2 poly-peptide chains of 32,000 and 34,000 dalton apparent molecular weight, which was expressed only on 25 +/- 10% of blood T lymphocytes and on 90 +/- 4% of thymus cells. Functional analysis of the T811+ and T811- T cell subsets isolated by rosetting with anti-mouse Ig coated ox erythrocytes revealed that both subpopulations were able to mount a proliferative response to allo-antigens, whereas allo-antigen induced cytotoxic cells and their precursors were only found in the T811+ subset. The pokeweed mitogen induced in vitro differentiation of B lymphocytes into immunoglobulin secreting cells was dependent on the presence of the T811- subset, whereas the T811+ T cells efficiently suppressed this differentiation.

Animals↗

Genetic specificity of primary and secondary proliferative and cytotoxic responses of human lymphocytes grown in continued culture.

Human T peripheral blood lymphocytes were grown in continued culture using conditioned medium obtained from phytohaemagglutinin-stimulated, pooled human leucocytes. These cultured T cells (CTC) were tested in mixed lymphocyte culture (MLC) and cell-mediated lympholysis (CML) assays to determine the genetic specificity of their proliferative and cytotoxic responses. Primary responses were measured after initial in vitro stimulation by allogeneic cells, and secondary responses were measured after a second in vitro stimulation by allogeneic cells. Both primary and secondary proliferative responses were found to be stimulated by alloantigens controlled by the HLA region and, more specifically, by antigens of the HLA-D region, in accordance with the responses of normal peripheral blood T lymphocytes. When CTC were established from unsensitized PBL and then stimulated with allogeneic cells, they could respond by proliferation in MLC, but, in contrast to PBL, they did not show subsequent cytotoxic responses. On the other hand, CTC established from PBL that had been stimulated first with allogeneic cells in either primary or secondary MLC displayed high levels of cytotoxic reactivity in CML. The strongest cytotoxicity was directed against allospecificities controlled by the HLA region and specific for the MLC-stimulating cells, but lower levels of cross-reactive cytotoxicity were also observed.

Cells, Cultured↗

Secondary MLC responses of primed lymphocytes after selective sensitization to non-HLA-D determinants.

Two HLA-B,D-identical siblings, who differed only for the HLA-A region because of a maternal recombinational event, were studied in primary (1 degrees) and secondary (2 degrees) mixed lymphocyte culture (MLC). The HLA-A:B recombinant child did not respond to its HLA-B,D-identical sibling in either 1 degrees or 2 degrees MLC. In the reciprocal combination the non-recombinant child responded only weakly in 1 degrees MLC but responded significantly in 2 degrees MLC to the HLA-A:B recombinant child. Thus, it was possible to selectively prime to a non-HLA-D determinant, which is controlled by a gene located distal to HLA-B. Because this determinant was not present on T-cells, it could be distinguished from the serologically defined antigen controlled by the HLA-A locus. Such primed lymphocytes, as well as lymphocytes primed between HLA-identical siblings, revealed high autologous control responses which were not observed when using lymphocytes primed in conventional one-haplotype combinations. The significant 2 degrees MLC response to autologous cells after sensitization to allogeneic cells may reflect recognition of self antigens and raises the question to what extent genetic similarity between responding and stimulating cells is required in the priming phase to elicit a 2 degrees response to autologous cells.

Child↗

Standardization of the human in vitro cell-mediated lympholysis technique.

Cytotoxic lymphocytes (CTL) can be generated in mixed lymphocyte culture and cryopreserved after a 6-day sensitization phase. CTL tested after storage in liquid nitrogen do not show lysis of autologous target cells and can mediate strong cytotoxicity against other target cells. This lysis shows the same specificity as that of the corresponding fresh CTL. The ability to cryopreserve CTL enables performance of sequential CML testing. Data from separate CML experiments can be pooled using a percent relative cytotoxic response (%RCR) to normalize values within individual experiments. Together, these methods provide a standardized cell-mediated lympholysis technique.

Cytotoxicity, Immunologic↗

The Qa-1 antigenic system. Relation of Qa-1 phenotypes to lymphocyte sets, mitogen responses, and immune functions.

The antiserum (B6 X A-Tlab) anti-A (Tlaa) defines several TL antigens expressed exclusively on thymocytes. When reacted with peripheral lymphocytes, the same antiserum defines another antigenic system, provisionally termed Qa-1. The genotypic disparity distinguishing the recipients and donors in this immunization comprises a section of chromosome 17 extending from a crossover point between H-2D and Tla to a presently unmarked point beyond Tla. Therefore although Qa-1 may constitute a single cell surface component, it is equally probable that the Qa-1 system defines two or more cell surface components determined by genes in this region, each of which may be expressed on a different cell set. Cytotoxicity assays indicate that Qa-1 antigen is expressed on Lyt-1 cells and Lyt-123 cells, and may serve to subclassify these two cell sets; it is not known whether Qa-1+ cells may occur within the small Lyt-23 set. There may be also be a cell set with the phenotype Thy-1--:Qa-1+. Another distinctive feature of the Qa-1 system is the characteristic profile of responses to mitogens exhibited by spleen cell populations from which Qa-1+ cells have been eliminated; in conventional assay of [3H]thymidine incorporation the response to lipopolysaccharide was essentially unchanged, the response to phytohemagglutinin M (PHA-M) was virtually abolished, and the response to concanavalin A (Con A) was reduced by 40%. The third distinctive feature of the Qa-1 system is the characteristic profile of changes which elimination of Qa-1+ cells produces in tests of immune function in vitro: (a) proliferation, measured by [3H]thymidine incorporation, in mixed lymphocyte culture (MLC) with major histocompatibility complex (MHC)-incompatible stimulator cells, was not affected. (b) in tests of cell-mediated cytotoxicity (CMC) of MHC-incompatible target cells, neither the generation nor the effector functions of cytotoxic lymphocytes was affected, implying that Lyt-23 prekiller and killer cells are Qa-1--. (c) primary and secondary responses to SRBC were considerably augmented, suggesting that Qa-1+ cells may be responsible for suppression in this test system. (d) accordingly the suppression of the anti-sheep erythrocyte (SRBC) response normally engendered in spleen cells by culture with SRBC was profoundly reduced by elimination of Qa-1+ cells, either before or after culture. (e) the suppression of the anti-SRBC response normally engendered in spleen cells cultured with Con A was reduced by removal of Qa-1+ cells before but not after culture with Con A. Although analysis is as yet far from complete, the Qa-1 system should already be of considerable value because it distinguishes a population of lymphocytes that is not defined by any other antigenic system, according to three criteria: (a) representation of Qa-1 cells among T-cell sets defined by Lyt phenotypes, (b) the profile of responses to mitogens exhibited by lymphocyte populations depleted of Qa-1+ cells, and (c) the profile of immune responses of lymphocyte populations depleted of Qa-1+ cells.

Animals↗

Cell-mediated lympholysis: examination of HLA genetic fine structure and complementation using cytotoxic lymphocytes.

Human cytotoxic lymphocytes (CTL), sensitized in vitro in one-way mixed lymphocyte cultures between two unrelated individuals, have been used to study target cell determinants. One pair of CTL shows strong cell-mediated lympholysis responses against the specific target cells as well as against third-party target cells obtained from unrelated individuals. Testing of sixty target cells shows a significant association between the two CTL with a correlation coefficient of 0.67 in the panel. The target determinant recognized by the CTL is not an HLA-A,B,C or D specificity, as defined by standard serological and cellular reagents. Segregation studies in a large family reveal that the new specificity defined by the CTL is controlled by HLA, and its recognition may be restricted by the HLA complex.

Cytotoxicity Tests, Immunologic↗