Biomedical subjects
Publications and source records attributed to H Waldmann.
Foam cells in 4 human atherosclerotic plaques reacted with 3 rat monoclonal antibodies directed against T.200 ('anti-leucocyte common' antibody), HLA-Class II molecules and macrophage cytoplasm, respectively. Smooth muscle cells did not react. The results support the view that foam cells are monocyte-derived macrophages.
Major histocompatibility complex (MHC) restrictions in the secondary antibody response in vitro to the soluble protein antigen trinitrophenyl keyhole limpet haemocyanin (TNP-KLH) were investigated. Experimental conditions were employed which ensured that co-operation between KLH-primed T cells and TNP-primed B cells was possible only through linked recognition of carrier and haptenic determinants. Under such conditions co-operation between T and B cells appears to be MHC-restricted. These experiments were designed to distinguish between two possible reasons for such MHC restrictions: (i) interaction between T and B cells is directly restricted by the MHC; or (ii) MHC-restricted T cell-antigen-presenting cell (APC) interactions limit generation of antibody responses and thus impose a pseudo-MHC restriction on T-B interactions. The ability of F1 APC to circumvent MHC restrictions was determined. The capacity of KLH-primed T cells from (P X Q)F1 leads to parent P and parent P leads to F1 X-irradiation bone marrow chimaeras to help TNP-primed parent P and parent Q B cells was assayed. Such T cells preferentially co-operate with parent P B cells. Addition of F1 APC from the spleen and the peritoneum did not alter the MHC restriction. Antibody responses stimulated by either high or low antigen concentrations were similarly MHC-restricted. Stimulation with antigen-pulsed F1 APC also failed to circumvent MHC restrictions. These results suggest that in this system involving linked recognition of antigenic determinants, the MHC directly restricts interactions between T and B cells.
Rapid segregation and purification of haemopoietic progenitor cells by simple methods is necessary for a proper analysis of the control of early haemopoiesis. In this paper we describe the use of a rat monoclonal antibody, YW 13.1.1, for that purpose. This reagent reacts with more than 90% of foetal liver cells but spares stem cells. A single-step lysis with antibody and complement achieves a tenfold enrichment for early progenitor cells. The marker also shows an increasing level of expression on the three defined subsets of erythroid progenitor cells. This parallels their developmental pathway and erythropoietin responsiveness. Simple quantitative considerations therefore permit separation of cells at different stages of erythropoiesis.
Mechanisms by which T and B lymphocytes co-operate in the in vitro secondary antibody response to trinitrophenyl (TNP)-conjugated soluble protein antigens were investigated. The generation of antibody responses was analyzed when haptenic and carrier determinants were either linked or non-linked. Ability to co-operate through each of these mechanisms was influenced by the experimental conditions employed, particularly the mode of preparation of the T cells and the antigen concentration used. Nylon wool filtration of T cells may deplete a T helper population involved in non-linked recognition interactions. High antigen concentrations favour the non-linked form of interaction whereas low antigen concentrations favour linked recognition interaction. These data suggest that at least two distinct co-operative mechanisms co-exist. However, experimental conditions can be defined under which either one mechanism predominates or where more than one mechanism is relevant.
From 1972 to 1981, 806 patients suffering from diseases of the bile ducts were seen at our Dept. 20% of them were over 70 years of age. Mortality rate, types of surgery, complications and causes of death are dealt with in detail. All the facts given demonstrate the necessity of early surgical intervention in case of surgical diseases of the bile ducts.
Nucleated cells from normal human peripheral blood and bone marrow have been analysed with 2 rat monoclonal antibodies of known specificity, one (YAML 555.6.6) directed against the P28,33 complex (anti-HLA-DR), and the other (YAML 501.4.4) directed against leucocyte common antigen (anti-LCA). The patterns of reactivity with an indirect immunoperoxidase method on fixed smeared cells were in close agreement with those obtained with a fluorescence activated cell sorter. A further new monoclonal antibody of unknown antigen specificity (YAML 537.2) reacts with an intracellular antigen present in neutrophils and their precursors from the promyelocyte stage onwards, megakaryocytes, and a proportion of monocytes, but not with eosinophils, nucleated red cells or lymphocytes. This reactivity could not be demonstrated using the fluorescence activated cell sorter. The immunoperoxidase method allows the identification of individual positive and negative cells and therefore provides a method of identifying minor reactive and non-reactive cell populations in a heterogeneous cell sample such as normal bone marrow. Cytoplasmic binding sites can be differentiated from membrane binding.
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Despite the range of available T cell specific monoclonal antibodies, there are no established rules to predict which might be immunosuppressive. We here describe a series of five rat monoclonal antibodies to a defined T cell antigen (mouse Thy-1) and evaluate their ability to immunosuppress mice. When compared with rabbit anti-lymphocyte globulin, only one of these monoclonal antibodies was able to delay skin allograft rejection and eliminate antibody responses to sheep red blood cells. This antibody was immunosuppressive following intra-peritoneal administration, even though it did not eliminate all of the T cells in vivo. Two factors may be relevant in determining the immunosuppressive properties of this reagent. First, the monoclonal antibody is of the rat IgG2b sub-class, and second, the specificity of the antibody is different to the other monoclonal antibodies in that it reacts with sub-populations of peripheral T cells, thymocytes and non-T cells. In practice, this suggests that to derive suitable monoclonal antibodies for human serotherapy, one should give attention to both the subclass and the fine specificity of the antibody for the target molecule.
Primitive hemopoietic progenitor cells from adult mouse marrow have been substantially enriched by virtue of a negative selection procedure with monoclonal antibodies. It has been possible to segregate erythroid progenitor cells at distinct stages of differentiation on the basis of their cell surface antigens. This has been achieved with two monoclonal antibodies reactive with the mature elements of bone marrow. YBM 34.3 binds to a heat-stable antigen expressed on B lymphocytes, neutrophils, and cells of the erythroid lineage. YBM 6.1 reacts with cells of the neutrophil, eosinophil, and monocyte series but does not bind to colony-forming cells. Separation is achieved by indirect immunoadsorption (panning) with YBM 34.3 on Protein-A-coated plastic plates followed by FACS II cell sorting with YBM 6.1. The combined procedures yield a marrow population containing 58% immature cells (blasts, promyelocytes, and myelocytes) and 9.5% clonogenic cells. In addition, differential binding of YBM 34.3 can be used to segregate erythroid progenitor cells at distinct stages of differentiation (day 7 BFU-E, day 5 BFU-E and CFU-E) either by cell sorting or panning. It is shown that both techniques give a comparable degree of resolution of the different cell types with, however, an appreciable advantage of panning over cell sorting in allowing the rapid handling of large numbers of cells.
Graft-versus-host disease is one of the major problems in clinical bone marrow transplantation. Many experiments in animals have shown that it could be greatly reduced if mature T lymphocytes were removed from the donor marrow. Here we describe a new rat monoclonal antibody, CAMPATH 1, which is suitable for depleting lymphocytes from human marrow grafts. CAMPATH 1 is an IgM that fixes human complement. It binds to both T and B lymphocytes and some monocytes but not to other hemopoietic cells. When peripheral blood mononuclear cells were treated with CAMPATH 1 and complement, more than 99% of lymphocytes were killed and viable T cells could no longer be detected. Under these conditions, in vitro multipotential erythroid and myeloid colony-forming cells were unaffected. As well as being used for in vitro treatment of bone marrow to remove T cells, CAMPATH 1 could potentially be applied to other experimental and clinical situations where depletion of lymphoid cells is required, including serotherapy to achieve immunosuppression for organ transplants or to treat lymphocytic leukemias.
We describe a family of six rat monoclonal antibodies which appear to recognize identical or closely related determinants present on virtually all mature human lymphocytes and monocytes, but absent from other blood cells including myeloid and erythroid colony-forming cells. Four IgM antibodies and one IgG2a fix human complement; one IgG2c antibody does so only poorly. All of the antibodies react with lymphocytes from baboons, rhesus and cynomolgus monkeys. However, in all baboons and rhesus monkeys tested, and some cynomolgus monkeys, they also recognize red cells. Nevertheless, in other cynomolgus monkeys reactivity is with lymphocytes only, so these animals will be suitable models for testing the effects of the antibodies in vivo. The antibodies described here could be useful for in vitro removal of lymphocytes from allogeneic marrow grafts (to prevent graft-versus-host disease) or malignant lymphoid cells from autologous grafts (for treatment of leukaemia) and may also find application as general immunosuppressants and for immunotherapy of leukaemia.
The safety and efficacy in vivo of three rat monoclonal anti-lymphocyte antibodies has been tested in cynomolgus monkeys. One IgM antibody, CAMPATH 1, was found to cause rapid but transient lymphopenia associated with consumption of complement. Two other antibodies with the same specificity, YTH 34.5 (IgG2a) and YTH 86.1 (IgG2c), had little effect. No acute or chronic toxic effects were associated with administration of any of the antibodies. CAMPATH 1 was used in a Phase I clinical trial for immunotherapy of two patients with terminal malignant lymphoid disease. It had no detectable toxic effects although it caused disappearance of circulating lymphocytes and consumption of complement. An inadequate rate of synthesis of complement components was a limitation to therapy in these patients with very large tumour burden, but might not be a major problem if antibody therapy were used at an earlier stage of the disease. CAMPATH 1 may also be useful as an immunosuppressive agent.
Erythropoietin-responsive progenitor cells (CFU-E) from normal mouse foetal liver have been substantially purified on the basis of their differential binding to two monoclonal antibodies in conjunction with flow cytometry. This has allowed the formal identification of the foetal liver CFU-E as an early erythroid blast cell with a highly basophilic cytoplasm. It has also been possible to show a quantitative association of a membrane marker with proliferative potential within a single differentiation lineage. This is the first demonstration of such an association. Progenitor cells enriched in this way should allow study of the molecular mechanisms of erythropoietin action and also serve as a target for determining the cellular specificity of erythroid-transforming viruses.
A solid-phase enzyme-linked binding assay is described for screening monoclonal antibodies to cell surface antigens. E. coli beta-galactosidase was coupled to rabbit anti-rat Ig and used to detect the binding of rat monoclonal antibodies to cells which had been fixed to the wells of microtitre plates using a combination of poly-L-lysine and glutaraldehyde. This method was found to be advantageous for the large screening of monoclonal antibodies with a panel of cell types, and has been useful in the selection of antibodies which would be candidates for differentiation markers within the human and mouse haemopoietic systems.
Monoclonal antibody (McAb) H9/25 has previously been shown to react with an alloantigen expressed on subpopulations of mouse lymphocytes. We here investigated the expression of the antigen (H9/25 Ag) on functional subsets of T and B cells. Lymphocytes were depleted of H9/25 Ag-bearing cells by complement-dependent cytolysis or by the affinity to immobilized McAb H9/25, and the residual immunological functions were tested. In these tests, killer T cells generated in mixed lymphocyte cultures and their precursors were found to express H9/25 Ag. In contrast, the activity and frequency of helper T cells specific to keyhole limpet hemocyanin carrier were not decreased by the depletion of H9/25 Ag-bearing cells. On the other hand, IgG and IgM anti-TNP plaque-forming cells were found to express H9/25 Ag, while it was not detected on unprimed B cells, as tested by using TNP-Ficoll and TNP-lipopolysaccharide. Only a small proportion of TNP memory B cells seemed to express the antigen. These results revealed significant differences between H9/25 Ag and other known alloantigens in the distribution among functional subsets of lymphocytes.
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Explore the source record for details and available documents.
Explore the source record for details and available documents.