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Biomedical subjects

D N Hart

Publications and source records attributed to D N Hart.

At least 91 records · Page 5Linked to original sources
PubMed1993

Expression of CD45 isoforms in chronic B-cell leukaemias.

CD45 isoform expression was studied in 204 cases of B-cell lymphoproliferative disease, including 162 chronic lymphocytic leukaemia (CLL). In almost half the samples tested, CD45R0 was co-expressed with CD45RA, unlike normal B-cells, which express only CD45RA, except at terminal stages of differentiation. In a small number of cases CD45R0 was the dominant isoform expressed. No correlation could be discerned either with Binet or RAI staging in CLL or with disease type CLL, prolymphocytic leukaemia (PLL) or hairy cell leukaemia (HCL). Comparison of CD45 isoform expression with other markers showed no correlation with apparent maturational status of the cells involved.

Antigens, CD↗
PubMed1993

Allostimulatory cells in fresh human blood: heterogeneity in antigen-presenting cell populations.

The relative ability of unmanipulated monocytes, B cells, and dendritic cells (DC) from peripheral blood to stimulate an allogeneic MLR has not been clearly established. We studied the allostimulatory ability of these cell types from minimally manipulated PBMC populations to exclude the induction of stimulatory properties by the complex isolation procedures commonly used to isolate blood DC. Highly purified cell populations were obtained from volunteer donors by immunolabeling PBMC with mAb directed against known lineage-associated markers and separating the positive and negative population on a FACS. These cells were used as stimulators in an allogeneic MLR. The major allostimulatory activity resides in the CD14, CD11b, and CD19 negative fractions. A mixture of antibodies to T, B, NK, monocyte, and FcRIII positive cells was then used to isolate a minor cell population that contained a markedly superstimulatory population of (CD3, CD14, CD16, CD19, and CD57) negative cells. We demonstrate that this activity is constitutive, and is not an artifact of the adherence and in vitro culture steps used in conventional DC purification procedures. We also show by rigorous depletion of the T cell responders that endogenous HLA class II positive cells in the responder population have little role in presenting processed allogeneic antigens during the primary MLR. Monocytes and B cells are stimulators of the allogeneic MLR, but are considerably less potent on a cell for cell basis than the putative DC population. Finally, because human blood and tonsil DC lack detectable CD43 by immunoperoxidase staining, in contrast to monocytes and activated B cells, we examined the ability of CD43 negative and positive cells to stimulate an allogeneic MLR. Similar allostimulatory activity for the human MLR was shown to reside in both the CD43 positive and negative fractions, suggesting that there may be some heterogeneity in the APC population.

Antigens, CD↗
PubMed1993

Dual CD45RA, CD45RO positive T-lymphocytes within rheumatoid arthritic joints.

This report documents the presence of an expanded population of dual CD45RA, CD45RO positive T cells (up to 91% of T cells) in the rheumatoid joint. Cells from peripheral blood (PB) and synovial fluid (SF) were analysed by dual immunofluorescence labelling. Synovium from a separate patient population was analysed by single and dual immunoenzyme staining of serial sections. Dual CD45RA, CD45RO positive T cells were found in PB (up to 74%), SF (up to 91%) and synovium. This was associated with a lack of early activation antigens (4F2, interleukin-2 receptor, transferrin receptor) but increased HLA-Class II antigens (HLA-DR, -DP, -DQ) in SF compared with PB. This intermediate activation phenotype may support the hypothesis that T cell activation or reactivation occurs within the rheumatoid joint.

Adult↗
PubMed1993

B7/BB-1 is a leucocyte differentiation antigen on human dendritic cells induced by activation.

Activation of a primary T-lymphocyte response requires additional signals apart from interaction of the T-cell receptor (TcR)/CD3 complex with major histocompatibility complex (MHC) antigens on the antigen-presenting cell. The CD28 antigen on T lymphocytes provides an important co-stimulatory signal to T lymphocytes and we therefore searched for the presence of its ligand, the B7/BB-1 antigen, on blood and tonsil dendritic cells (DC). Blood DC, prepared from peripheral blood mononuclear cells with a minimal period of in vitro culture, did not stain with the monoclonal antibody BB-1 using flow cytometry analysis. In contrast, tonsil DC stained weakly for B7/BB-1 compared to positive control cell lines. Polymerase chain reaction (PCR) was used to amplify a 605 base pair (bp) fragment from human B7/BB-1 mRNA and demonstrated significant amounts of B7/BB-1 mRNA in tonsil DC but no specific product was obtained from blood DC, confirming the surface-staining results. Weak expression of B7/BB-1 antigen was detected by immunofluorescence analysis following culture of blood DC with either interferon-gamma (IFN-gamma) or granulocyte-macrophage colony-stimulating factor (GM-CSF). These data support the concept that blood DC give rise to tissue and/or lymphoid DC, which acquire co-stimulatory ligands as a result of activation and/or differentiation.

Antibodies, Monoclonal↗
PubMed1993

Expression of the CMRF-35 antigen, a new member of the immunoglobulin gene superfamily, is differentially regulated on leucocytes.

A new monoclonal antibody, CMRF-35, has been generated that recognized a 224 amino acid cell surface protein which is a novel member of the immunoglobulin gene superfamily. The antibody, raised against large granular lymphocytes (LGL), stains LGL, monocytes, macrophages and granulocytes but not platelets or erythrocytes. In addition, a subset of peripheral blood T lymphocytes (26.6 +/- 13.4% CD5+ cells) and B lymphocytes (13.7 +/- 6.8% CD20+ cells) stained with CMRF-35 but tonsil T and B cells were essentially negative. Expression of the CMRF-35 antigen (Ag) on different leucocyte populations was markedly influenced by stimulation of the cells with mitogens and cytokines. Activation of peripheral blood T cells with phytohaemagglutinin (PHA), or phorbol myristate acetate (PMA) and calcium ionophore (CaI) led to a decrease in the proportion of CMRF-35+ T lymphocytes. In contrast, PHA activation of tonsil T lymphocytes resulted in an increase in CMRF-35 Ag expression (47.1 +/- 1.5% CD5 cells at 6 days). An increase in CMRF-35 Ag was also seen on phorbol ester and CaI-activated tonsil B cells. No change in CMRF-35 expression on natural killer (NK) cells occurred following activation with interleukin-2 (IL-2) but the CMRF-35 Ag was down-regulated following Fc receptor stimulation. A moderate increase in CMRF-35 expression occurred during monocyte-macrophage differentiation and the expression of the Ag on monocytes was differentially regulated by interferon-gamma (IFN-gamma). This regulation of the CMRF-35 Ag on the leucocyte surface suggests that the molecule has an important function common to diverse leucocyte types.

Antibodies, Monoclonal↗
PubMed1992

Epstein-Barr viral DNA in acute large granular lymphocyte (natural killer) leukemic cells.

Serologic studies in a male Caucasian presenting with an acute hepatitis-like illness, associated with an increase in peripheral blood large granular lymphocytes (LGLs), suggested a chronic or reactived Epstein-Barr virus (EBV) infection. The LGL were shown to have a natural killer (NK) cell, CD3- CD16- CD56+ CD57- phenotype and mediated strong nonspecific major histocompatability complex-unrestricted (NK) cytotoxic activity. A progressive increase in the peripheral blood LGL count was associated with a rapid deterioration, hepatic necrosis, and death. Widespread organ infiltration with LGLs suggested a malignant lymphoproliferative condition, but no lymphoid (T-cell receptor or IgH) gene rearrangement or cytogenetic marker was detected. However, molecular analysis identified EBV genomic DNA present in a single episomal form within the LGL, establishing the clonal nature of the LGL proliferation. Confirmation that the EBV had infected the leukemic LGL was obtained by in situ hybridization studies that showed EBV RNA within the LGLs. Immunoblotting of LGL protein extracts established that, of the EBV gene products, EBV nuclear antigen-1 (EBNA-1) was expressed but EBNA-2 and the latent membrane protein (LMP-1) were not detectable in the leukemic cells. These results suggest that EBV may be involved directly in LGL cell transformation, in a manner similar to EBV-associated B-cell lymphomas, although other molecular changes probably contribute to the evolution of a fully malignant leukemic clone.

Antigens, CD↗
PubMed1992

Molecular cloning of a novel member of the immunoglobulin gene superfamily homologous to the polymeric immunoglobulin receptor.

The CMRF35 monoclonal antibody recognizes a cell membrane antigen present on the surface of monocytes, neutrophils, a proportion of peripheral blood T and B lymphocytes and lymphocytic cell lines. Initial studies with CMRF35 suggested an unusual pattern of serological reactivity which did not correspond to any of the known leukocyte differentiation antigen clusters. We describe here the cloning and sequencing of a cDNA encoding the CMRF35 antigen by means of expression in COS cells and immunoselection with the CMRF35 monoclonal antibody. The cDNA encodes a novel integral membrane glycoprotein of 224 amino acids that represents a new member of the immunoglobulin (Ig) gene superfamily. The molecule comprises (a) a single extracellular Ig variable domain remarkably similar to the Fc receptor for polymeric IgA and IgM, (b) a membrane-proximal domain containing a high proportion of proline, serine and threonine residues that was predicted to be heavily O-glycosylated, (c) an unusual transmembrane anchor that contained a glutamic acid and a proline residue and (d) a short cytoplasmic tail. Transcripts encoding the CMRF35 protein were detected in early monocytic cell lines, in peripheral blood T cells and in some B lymphoblastoid cell lines, confirming the results of immunocytological staining. However, the level of CMRF35 expression on peripheral blood T cells was shown to decrease in response to mitogenic stimulation. The likelihood that the CMRF35 antigen shares a common evolutionary ancestor with the poly Ig Fc receptor is discussed.

Amino Acid Sequence↗
PubMed1992

Cellular protein profiles of the Hodgkin's disease cell lines L428, KM-H2 and HDLM-2: a comparative study.

The origin of the malignant mononuclear Hodgkin's cell and the classic Reed-Sternberg cell in Hodgkin's disease (HD) remains controversial despite extensive immunohistological and lymphoid gene analysis. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) in combination with protein staining and radioactive labelling has not been fully exploited in analysing the HD-derived cell lines available. The NP40 solubilised cellular proteins from the three HD cell lines, L428, KM-H2 and HDLM-2 were analysed using these techniques and compared with other haemopoietic cell lines and leucocytes of myeloid and lymphoid origin. The electrophoretic patterns of the three HD cell lines, although clearly different from one another, had many features in common. No major differences between the cell types were detected by Coomassie brilliant blue staining. The HD cell lines were more readily distinguished from the myeloid and to a lesser extent the lymphoid cell lines by silver staining, but HD cell line specific proteins (13, 19, 36, 60, 150 kD) were detected only on one line, L428. Iodination of cell membrane molecules, SDS-PAGE and subsequent autoradiography revealed three molecules (118, 22, 12 kD) which were restricted to the HD cell lines and the B-cell line Mann, and one molecule (144 kD) restricted to the HD cell lines and U937. Molecules unique to HDLM-2 (211 kD) and L428 (46 kD) were also detected by this method. Cell surface labelling with NaB3H4 identified a glycoprotein of 102 kD limited to HDLM-2 and L428, as well as a glycoprotein of 97 kD present on KM-H2 alone and one of 63 kD on L428 alone. Overall the HD cell line protein profiles displayed little similarity to the patterns of the other cell types studied and provide further evidence to support functional and phenotypic studies which identify Hodgkin's cells as a unique cell type. The molecules identified as HD cell line restricted may have potential as markers for this cell type.

Cell Membrane↗
PubMed1992

Adhesion molecules on human tonsil dendritic cells.

Dendritic cells are specialist antigen-presenting cells that have a unique ability to stimulate a primary T cell response. Activation of T cells by DC depends on the formation of cell clusters creating DC-T cell membrane contact that probably involves adhesion molecules. Monoclonal antibodies were used to study adhesion molecules on DC, including members of the integrin and immunoglobulin supergene families. DC expressed LFA-1, ICAM-1, LFA-3, and the Hermes antigen, but no other integrin or immunoglobulin supergene family adhesion molecules were detected using a sensitive immunoperoxidase staining technique. Monoclonal antibodies to LFA-1 alpha and LFA-1 beta inhibited DC-stimulated allogeneic T cell (MLR) responses by 75 +/- 12% and 74 +/- 8%, respectively, as did the anti-LFA-3 (56 +/- 3% inhibition) and anti-LFA-2 (60 +/- 5% inhibition) antibodies. Three different anti-ICAM-1 antibodies inhibited only to a limited degree (mean range 8-24%). The inhibitory effect of the LFA-1 and LFA-3 antibodies was maximal if added early to the MLR. The inhibitory effect of the different antibodies was associated with variable decreases in DC-T cell cluster stability. The simultaneous addition of monoclonal antibodies to MLRs and preincubation washing experiments established that DC have at least 3 independent adhesion ligand interactions (LFA-1-ICAM-1, ICAM-1-LFA-1, and LFA-3-CD2) with T cells. It seems likely that the additional ligand for LFA-1, ICAM-2, is expressed on DC and contributes significantly to DC-T cell adherence and T cell activation. The membrane mobility of these molecules may also be important in the DC-T cell activation process.

Antibodies↗
PubMed1992

Hodgkin's cells express a novel pattern of adhesion molecules.

Adhesion molecules play an important role in the functioning of the immune system, particularly with regard to cell-cell interactions and antigen presentation. Several adhesion molecules are expressed on Hodgkin's disease-derived cell lines and these are important in their molecular interactions as antigen presenting cells (APC). There are no data regarding the expression of many of these adhesion molecules on Reed-Sternberg cells and its mononuclear variant (Hodgkin's cells (HC)) present in pathological material. To obtain this information we undertook an immunohistological study on material from 18 cases of Hodgkin's disease using a panel of MoAbs to examine the expression of adhesion molecules on HC. The HC were shown to express the integrin beta 1 subfamily molecules, LFA-1 (CD11a) and p150,95 (CD11c) in high density but lacked CR3 (CD11b). All of the immunoglobulin gene superfamily adhesion molecules studied were present to some degree on HC, with ICAM-2, in particular, showing moderate to strong expression in most cases. The Hermes antigen CD44 was present in high density but leukosialin (CD43), another molecule present on diverse leucocyte types, was, in general, not detected on HC. These new data showing that ICAM-1, ICAM-2 and LFA-3 are, like LFA-1, expressed on HC emphasize the ability of HC to act as APC. The known adhesion molecule phenotype of the recently defined haematopoietic lineage of human dendritic cells (DC) is broadly similar to that of HC, perhaps supporting the hypothesis that some HC represent a malignancy of an APC (DC) lineage.

Antibodies, Monoclonal↗
PubMed1992

Hodgkin's disease cell lines: a model for interleukin-1-independent accessory cell function.

The haemopoietic origins of the Hodgkin's disease (HD)-derived cell lines L428, KM-H2 and HDLM-2 remain controversial. Analysis of T-cell receptor (TcR) and Ig rearrangements cannot resolve this, and lineage promiscuity limits the interpretation of isolated surface antigen expression. Nonetheless the cell marker profile of L428 has similarities with human dendritic cells (DC), and L428 strongly stimulates in the mixed leucocyte reaction (MLR). We therefore undertook an extended immunophenotypic comparison of the HD lines with that recently defined for DC, prior to examining their ability to stimulate allogenic T lymphocytes, and comparing the molecular interactions involved with those of primary MLR stimulatory cells. The immunophenotype of the HD lines failed to establish either a lymphoid or monocytoid derivation. The profile of L428 appeared similar to the human DC. All three lines were potent stimulators in the primary MLR, and each expressed relevant adhesion and signal-transducing molecules important for co-stimulating T lymphocytes. Inhibition studies using monoclonal antibodies indicated similar contributions within HD line-T cell MLR to that documented in human tonsil DC-T cell MLR. The HD lines produced no detectable interleukin-1 (IL-1) by biological or immunological analysis. Moreover they stimulated allogeneic T lymphocytes in the presence of anti-IL-1 antibodies. Thus although IL-1 mRNA can be detected in both HDLM-2 and KM-H2 by polymerase chain reaction, these lines, and L428, share with DC the ability to stimulate allogeneic T lymphocytes in an IL-1-independent manner [corrected]. HD lines, particularly L428, may provide a standardized, reproducible, IL-1-independent model for dissection of the co-stimulatory requirements of the human primary MLR.

Antigens, CD↗
PubMed1991

The costs and benefits of bone marrow transplantation.

The average direct costs of performing a bone marrow transplant (BMT), including the subsequent year, was found to be NZ$27,074 for 43 consecutive transplants. In 29 BMTs a full two year period of follow up was available and a quality of life analysis was carried out on these patients. It was calculated that 59 quality adjusted life years (QALYs) had been gained by the BMT procedure at the time of analysis. By combining these two analyses the cost of each QALY gained by BMT is NZ$13,272. The relatively low cost of BMT is partly due to the extremely low annual costs in second and subsequent years post BMT. In our patients this cost amounted to $195 per year. The costs and benefits of BMT compare very favourably with other complex medical procedures.

Adult↗
PubMed1991

Intranasal amphotericin B reduces the frequency of invasive aspergillosis in neutropenic patients.

PURPOSE: To retrospectively study the prophylaxis of invasive aspergillosis in neutropenic patients and to relate the frequency of this fungal disease to any causal or modifying factors that could be identified. PATIENTS AND METHODS: Between 1977 and 1988, 130 patients underwent 158 intensive treatment episodes to control acute leukemia, lymphoma, and aplastic anemia, and the frequency of complicating aspergillus infection was determined. RESULTS: Proven invasive aspergillus infections occurred in 22 cases, 12 of which were fatal. Invasive aspergillosis was suspected in a further 16 cases and all these patients recovered with amphotericin B treatment. Colonization by Aspergillus in the absence of clinically significant infection was seen in 31 treatment episodes. Invasive aspergillosis involved mainly the upper and lower respiratory tract and skin. Control of the infection was closely related to the control of the underlying disease, with subsequent return of normal marrow function and resolution of neutropenia. The incidence of aspergillus infection has decreased dramatically since 1985, most probably due to the introduction of intranasal amphotericin B. This occurred despite the persistence of aspergillus spores in the hematology ward air during the 1986 to 1988 period. CONCLUSION: Intranasal aerosolized amphotericin B may protect against invasive aspergillosis, even when neutropenic patients are cared for in conventional wards without HEPA filtration.

Administration, Intranasal↗