D N Hart

Canterbury District Health Board, Christchurch, Canterbury Region, New Zealand

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Publications (122)439.78 Total impact

  • 01/2011;
  • Transplantation 01/2008; 86. · 3.78 Impact Factor
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    ABSTRACT: To facilitate human multiple myeloma (MM) engraftment into NOD/SCID recipients, mice were depleted of CD122+ cells (NK and myeloid cells) by antibody-mediated ablation prior to transplantation with the MM cell lines (RPMI8226, RPMI8226-TGL or U226). The MM engraftment, skeletal MM distribution, osteolysis, lambda chain paraprotein and associating disease symptoms in CD122+ cell-depleted and CD122+ cellreplete mice were compared. The CD122+ cell-depleted mice engrafted at a significantly higher frequency with human CD38+, CD56+, PCA-1+ and CD138+ cells. In the CD122+ cell-depleted mice, bioluminescence MM signal involved the whole mouse compared to limited imaging signal in the CD122+ cell-replete mice. The majority 88%–100% of CD122+ cell-depleted mice developed MM engraftment throughout the appendicular and axial skeletons with osteolysis and rare subcutaneous plasmacytomas (11% of mice). Serum paraprotein appeared earlier at 4–5 weeks post-transplant in CD122+ cell-depleted mice and continued to increase during the 12–13 weeks of analysis. The majority, 92% of CD122+ cell-depleted mice developed hind-limb paralysis and had a significantly shortened 45 day survival. Thus, depletion of CD122+ cells reduced resistance to the human MM and produced a new MM xenograft-NOD/SCID model that recapitulates the clinical manifestations of MM and eliminates the major limitations associated with the published MM xenograft-mouse model.
    Blood 01/2008; 112:2734. · 9.78 Impact Factor
  • Transplantation 01/2008; 86. · 3.78 Impact Factor
  • Source
    K. L. SUMMERS, J. L. O'DONNELL, D. N. J. HART
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    ABSTRACT: The site of T lymphocyte activation in chronic arthritis is unknown. Peripheral blood (PB) lymphocytes from chronic arthritis patients are in a ‘naïve’ or non-activated state, as defined by expression of the CD45RA antigen and lack of HLA class II expression. In contrast, most synovial fluid (SF) T lymphocytes express a ‘memory’ or activated phenotype, as defined by the CD45RO antigen and high HLA class II expression. Following stimulation, naive cells lose CD45RA and gain CD45RO expression to become memory cells with a transitional stage of dual CD45RA, CD45RO antigen expression. To localize where this change in phenotype occurs we used dual colour immunofluorescence labelling to compare the percentage of dual CD45RA, CD45ROpositive T lymphocytes in PB and SF from chronic arthritic patients and from normal PB, assuming this population would be increased at the primary site of T lymphocyte activation. Expression of the intermediate and late activation marker. HLA-DR, was also analysed using dual colour immunofluorescence labelling. The percentage of dual positive T lymphocytes was similar between arthritic PB, SF. and normal PB, as was the density of both CD45RA and CD45RO antigens. Thus, CD45 isoform expression did not indicate where T lymphocytes were activated. However, we identified a previously unreported population of CD45RA+ CD45RO+ HLA-DR- T lymphocytes in arthritic and normal PB. In SF, this population was absent, but a substantial number of dual CD45RA, CD45RO-positive HLA-DR+ T lymphocytes were identified. This population would not be predicted by the current model of T lymphocyte activation. Division of T lymphocytes into functional groups on the basis of CD45 isoform expression is likely to be more complicated than previously thought. Based on our findings we propose an alternative model of T lymphocyte differentiation.
    Clinical & Experimental Immunology 01/2008; 97(1):39-44. · 3.41 Impact Factor
  • Biology of Blood and Marrow Transplantation - BIOL BLOOD MARROW TRANSPLANT. 01/2006; 12(2):12-13.
  • Biology of Blood and Marrow Transplantation - BIOL BLOOD MARROW TRANSPLANT. 01/2006; 12(2):119-119.
  • D N Hart
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    ABSTRACT: Dendritic cells (DC) are now recognised as a unique leukocyte type, consisting of two or more subsets. The origins and functional inter-relationships of these cells are the subject of intense basic scientific investigation. They play important roles in initiating and directing immune responses, defending the host from pathogens and maintaining self tolerance. Fundamental studies are defining new molecules and mechanisms associated with DC function. The first methods for counting these rare blood cell populations are already providing interesting new clinical data. Indeed, abnormal DC function may contribute to deficiencies in the immune response against malignancies. Phase I trial data suggests that DC-based cancer vaccination protocols may contribute an important new biological approach to cancer therapy. Manipulation of DC to facilitate allogeneic transplantation and even to manage autoimmune disease are likely developments.
    Pathology 12/2001; 33(4):479-92. · 2.66 Impact Factor
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    ABSTRACT: Dendritic cells (DC) are the only antigen-presenting cells that can activate naïve T lymphocytes and initiate a primary immune response. They are also thought to have a role in immune tolerance. DC traffic from the blood to peripheral tissue where they become activated. They then present antigen and the costimulating signals necessary to initiate an immune response. In this study, we investigated the number, subsets, and activation pattern of circulating and intestinal DC from patients with clinically mild ulcerative colitis (UC) or Crohn's disease. Patients were recruited, if they were not taking immunosuppressive therapy, and were assessed for clinical severity of their disease using for UC, the Clinical Activity Index, and for Crohn's disease, the Crohn's Disease Activity Index. Blood CD11c+ and CD11c- DC subsets, expression of costimulatory antigens, CD86 and CD40, and the early differentiation/activation antigen, CMRF44, were enumerated by multicolor flow cytometry of lineage negative (lin- = CD3-, CD19-, CD14-, CD16-) HLA-DR+ DC. These data were compared with age-matched healthy and the disease control groups of chronic noninflammatory GI diseases (cGI), acute noninflammatory GI diseases (aGI), and chronic non-GI inflammation (non-GI). In addition, cryostat sections of colonoscopic biopsies from healthy control patients and inflamed versus noninflamed gut mucosa of inflammatory bowel disease (IBD) patients were examined for CD86+ and CD40+ lin- cells. Twenty-one Crohn's disease and 25 UC patients, with mean Crohn's Disease Activity Index of 98 and Clinical Activity Index of 3.1, and 56 healthy controls, five cGI, five aGI, and six non-GI were studied. CD11c+ and CD11c- DC subsets did not differ significantly between Crohn's, UC, and healthy control groups. Expression of CD86 and CD40 on freshly isolated blood DC from Crohn's patients appeared higher (16.6%, 31%) and was significantly higher in UC (26.6%, 46.3%) versus healthy controls (5.5%, 25%) (p = 0.004, p = 0.012) and non-GI controls (10.2%, 22.8%) (p = 0.012, p = 0.008), but not versus cGI or aGI controls. CD86+ and CD40+ DC were also present in inflamed colonic and ileal mucosa from UC and Crohn's patients but not in noninflamed IBD mucosa or normal mucosa. Expression of the CMRF44 antigen was low on freshly isolated DC, but it was upregulated after 24-h culture on DC from all groups, although significantly less so on DC from UC versus Crohn's or healthy controls (p = 0.024). The CMRF44+ antigen was mainly associated with CD11c+ DC, and in UC was inversely related to the Clinical Activity Index (r = -0.69, p = 0.0002). There is upregulation of costimulatory molecules on blood DC even in very mild IBD but surprisingly, there is divergent expression of the differentiation/activation CMRF44 antigen. Upregulation of costimulatory molecules and divergent expression of CMRF44 in blood DC was also apparent in cGI and aGI but not in non-GI or healthy controls, whereas intestinal CD86+ and CD40+ DC were found only in inflamed mucosa from IBD patients. Persistent or distorted activation of blood DC or divergent regulation of costimulatory and activation antigens may have important implications for gut mucosal immunity and inflammation.
    The American Journal of Gastroenterology 11/2001; 96(10):2946-56. · 9.21 Impact Factor
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    ABSTRACT: CD83 is an inducible glycoprotein expressed predominantly by dendritic cells (DC) and B lymphocytes. Expression of membrane CD83 (mCD83) is widely used as a marker of differentiated/activated DC but its function and ligand(s) are presently unknown. We report the existence of a soluble form of CD83 (sCD83). Using both a sCD83-specific ELISA and Western blotting, we could demonstrate the release of sCD83 by mCD83(+) B cell and Hodgkin's disease-derived cell lines, but not mCD83(-) cells. Inhibition of de novo protein synthesis did not affect the release of sCD83 during short-term (2 h) culture of cell lines although mCD83 expression was significantly reduced, suggesting sCD83 is generated by the release of mCD83. Isolated tonsillar B lymphocytes and monocyte-derived DC, which are mCD83(low), released only low levels of sCD83 during culture. However, the differentiation/activation of these populations both up-regulated mCD83 and increased sCD83 release significantly. Analysis of sera from normal donors demonstrated the presence of low levels (121 +/- 3.6 pg/ml) of circulating sCD83. Further studies utilizing purified sCD83 and the analysis of sCD83 levels in disease may provide clues to the function and ligand(s) of CD83.
    International Immunology 08/2001; 13(7):959-67. · 3.14 Impact Factor
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    ABSTRACT: Dendritic cells (DCs) are specialized antigen-presenting cells that have the unique ability to initiate a primary immune response. The effect of physiologic stress on circulating blood DCs has thus far not been studied. In this study, we applied a recently developed method of counting blood DCs to test the hypothesis that significant stress to the body such as surgery and exercise might induce measurable changes in the DC numbers, subsets, phenotype, and function. Twenty-six patients scheduled for elective laparoscopic cholecystectomy, 4 for elective hysterectomy, 56 controls, and 5 volunteers who underwent a stress exercise test were enrolled in the study. Absolute DC counts increased acutely (71.7% +/- 11% [SEM], P =.0001) in response to the stress of surgery and dropped below preoperative levels (-25% +/- 14% [SEM], P =.05) on days 2-3. The perioperative DC subset balance remained constant. Interestingly, DC counts changed independently of monocyte counts. Exercise also induced a rise in DC counts but coincidentally with monocyte counts. Surprisingly, no phenotypic or functional activation of DCs was seen in either stress situations in vivo. DCs are rapidly mobilized into the circulation in response to surgical and exercise stress, which may serve to prepare the host's immune defenses against trauma. The independent regulation of the DC and monocyte counts reinforces the distinction between these 2 cell populations.
    Blood 08/2001; 98(1):140-5. · 9.78 Impact Factor
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    ABSTRACT: As dendritic cells increasingly become the adjuvant of choice in new approaches to cancer immunotherapy, a degree of protocol standardization is required to aid future large-scale clinical trials.
    Nature Medicine 08/2001; 7(7):761-5. · 22.86 Impact Factor
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    ABSTRACT: Heterogeneous expression of several antigens on the three currently defined tonsil dendritic cell (DC) subsets encouraged us to re-examine tonsil DCs using a new method that minimized DC differentiation and activation during their preparation. Three-color flow cytometry and dual-color immunohistology was used in conjunction with an extensive panel of antibodies to relevant DC-related antigens to analyze lin(-) HLA-DR(+) tonsil DCs. Here we identify, quantify, and locate five tonsil DC subsets based on their relative expression of the HLA-DR, CD11c, CD13, and CD123 antigens. In situ localization identified four of these DC subsets as distinct interdigitating DC populations. These included three new interdigitating DC subsets defined as HLA-DR(hi) CD11c(+) DCs, HLA-DR(mod) CD11c(+) CD13(+) DCs, and HLA-DR(mod) CD11c(-) CD123(-) DCs, as well as the plasmacytoid DCs (HLA-DR(mod) CD11c(-) CD123(+)). These subsets differed in their expression of DC-associated differentiation/activation antigens and co-stimulator molecules including CD83, CMRF-44, CMRF-56, 2-7, CD86, and 4-1BB ligand. The fifth HLA-DR(mod) CD11c(+) DC subset was identified as germinal center DCs, but contrary to previous reports they are redefined as lacking the CD13 antigen. The definition and extensive phenotypic analysis of these five DC subsets in human tonsil extends our understanding of the complexity of DC biology.
    American Journal Of Pathology 08/2001; 159(1):285-95. · 4.60 Impact Factor
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    ABSTRACT: The immunoregulatory signaling (IRS) family includes several molecules, which play major roles in the regulation of the immune response. The CMRF-35A and CMRF-35H molecules are two new members of the IRS family of molecules, that are found on a wide variety of haemopoietic lineages. The extracellular functional interactions of these molecules is presently unknown, although CMRF-35H can initiate an inhibitory signal and is internalized when cross-linked. In this paper, we described the gene structure for the CMRF-35A gene and its localization to human chromosome 17. The gene consists of four exons spanning approximately 4.5 kb. Exon 1 encodes the 5' untranslated region and leader sequence, exon 2 encodes the immunoglobulin (Ig)-like domain, exon 3 encodes the membrane proximal region and exon 4 encodes the transmembrane region, the cytoplasmic tail and the 3' untranslated region. A region in the 5' flanking sequence of the CMRF-35A gene, that promoted expression of a reporter gene was identified. The genes for the CMRF-35A and CMRF-35H molecules are closely linked on chromosome 17. Similarity between the Ig-like exons and the preceding intron of the two genes suggests exon duplication was involved in their evolution. We also identified a further member of the CMRF-35 family, the CMRF-35J pseudogene. This gene appears to have arisen by gene duplication of the CMRF-35A gene. These three loci - the CMRF-35A, CMRF-35J and CMRF-35H genes-form a new complex of IRS genes on chromosome 17.
    Tissue Antigens 06/2001; 57(5):415-23. · 2.93 Impact Factor
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    ABSTRACT: Dendritic cells (DC) are potent antigen-presenting cells and understanding their mechanisms of antigen uptake is important for loading DC with antigen for immunotherapy. The multilectin receptors, DEC-205 and macrophage mannose receptor (MMR), are potential antigen-uptake receptors; therefore, we examined their expression and FITC-dextran uptake by various human DC preparations. The RT-PCR analysis detected low levels of DEC-205 mRNA in immature blood DC, Langerhans cells (LC) and immature monocyte-derived DC (Mo-DC). Its mRNA expression increased markedly upon activation, indicating that DEC-205 is an activation-associated molecule. In Mo-DC, the expression of cell-surface DEC-205 increased markedly during maturation. In blood DC, however, the cell-surface expression of DEC-205 did not change during activation, suggesting the presence of a large intracellular pool of DEC-205 or post-transcriptional regulation. Immature Mo-DC expressed abundant MMR, but its expression diminished upon maturation. Blood DC and LC did not express detectable levels of the MMR. FITC-dextran uptake by both immature and activated blood DC was 30- to 70-fold less than that of LC, immature Mo-DC and macrophages. In contrast to immature Mo-DC, the FITC-dextran uptake by LC was not inhibited effectively by mannose, an inhibitor for MMR-mediated FITC-dextran uptake. Thus, unlike Mo-DC, blood DC and LC do not use the MMR for carbohydrate-conjugated antigen uptake and alternative receptors may yet be defined on these DC. Therefore, DEC-205 may have a different specificity as an antigen uptake receptor or contribute to an alternative DC function.
    International Immunology 12/2000; 12(11):1511-9. · 3.14 Impact Factor
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    ABSTRACT: Cardiac failure is a known complication of haemopoietic stem cell transplantation (HSCT) and is often difficult to diagnose as patients may have multiple medical problems. Since brain natriuretic peptide (BNP) is largely a hormone of cardiac ventricular origin and is released early in the course of ventricular dysfunction, we have examined the value of serial plasma BNP levels for detecting cardiac failure in patients undergoing cytotoxic conditioning for HSCT. Fifteen patients undergoing HSCT were evaluated (10 undergoing autologous HSCT; five undergoing allogeneic HSCT). BNP was measured by radioimmunoassay prior to therapy and weekly for 5 weeks. Seven patients had a significant rise in BNP level (above a previously established threshold of 43 pmol/l associated with cardiac failure), occurring 1-4 weeks post commencement of conditioning. In three of these patients, cardiac failure was subsequently diagnosed clinically 3, 9 and 23 days after a BNP level of 43 pmol/l had been detected. These three patients had the highest peak BNP levels for the group and in each case elevation in BNP level occurred for a period exceeding 1 week. Although numbers were relatively small, a BNP >43 pmol/l was significantly associated with the inclusion of high-dose cyclophosphamide in the preparative regimen (P = 0.02). BNP levels showed no relationship to febrile episodes. In conclusion, these results show that plasma BNP may be used as a marker for early detection of cardiac dysfunction in patients undergoing HSCT, particularly if levels are increased for periods exceeding 1 week. Measurement of BNP during HSCT may be helpful in patients at risk of cardiac failure, in complex clinical situations and in monitoring the cardiotoxicity of preparative regimens.
    Bone Marrow Transplantation 08/2000; 26(3):309-13. · 3.54 Impact Factor
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    ABSTRACT: We describe a man with relapsed large B cell mediastinal lymphoma and associated infected large anterior chest wall defect who required high dose salvage therapy for his underlying disease. An initial mediastinotomy wound, associated with recurrent sepsis, had developed into an abscess, then fistula and eventually a large anterior chest wall defect. Safe use of salvage chemotherapy required reconstructive surgery consisting of a pedicled muscle flap. The subsequent high dose chemotherapy was carried out without complications and 15 months later the patient is alive and well.
    Clinical & Laboratory Haematology 05/2000; 22(2):127-8. · 1.11 Impact Factor
  • G J Clark, B J Green, D N Hart
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    ABSTRACT: The CMRF-35 monoclonal antibody recognizes an epitope found on at least two cell surface molecules, differentially expressed by many leukocytes. These molecules, the CMRF-35H (9) and CMRF-35A (CMRF-35) antigens are both members of the immunoglobulin (Ig) superfamily with a single V-like Ig domain. The function of these molecules is unknown, however the presence of putative immunoreceptor tyrosine-based inhibitory motifs (ITIM) in the cytoplasmic domain of the CMRF-35H molecule suggests that this molecule may play a regulatory role in leukocyte function. The CMRF-35H and CMRF-35A molecules show several similarities to the family of molecules containing ITIM or immunoreceptor tyrosine-based activatory motifs (ITAM) suggesting that CMRF-35H/CMRF-35A may be new members of this family. This would further indicate that, like other ITIM/ITAM containing molecules, CMRF-35H/CMRF-35A will also play an important role in the immune response. To further characterize these molecules, we have isolated genomic clones for the CMRF-35H gene and determined its intron-exon organization. The gene spans approximately 12 kb and consists of seven exons. Furthermore, this gene has been mapped to chromosome 17 and thus is not linked to the known human ITIM containing genes which map to human chromosome 19 or the recently characterized molecule, NKp44, localized to human chromosome 6.
    Tissue Antigens 03/2000; 55(2):101-9. · 2.93 Impact Factor
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    ABSTRACT: To determine if dendritic antigen-presenting cells (DC) are present in rheumatoid nodules, as has been reported in the synovial lesions of rheumatoid arthritis. Nodules (n = 14) were examined with monoclonal antibodies (Mab) recognizing the DC differentiation/activation markers CD83, CMRF44, and CMRF56 and an antibody recognizing the CD1a antigen present on epithelial tissue associated DC. Results. Cells expressing CMRF44 were common in rheumatoid nodules, comprising 22% of nucleated cells versus 13% in synovial membranes (n = 10). Cells positive for CD1a (5%) and CD83 (2%) were less common. A majority (86%) of CMRF44 positive cells were also positive for the macrophage marker CD14. This left a significant minority of putative DC that were single stained with CMRF44. Cells bearing DC markers are as frequent in the rheumatoid nodule as in the synovial lesions. A majority are "indeterminate" cells that are CD14 positive but a proportion are single stained putative DC. The lack of lymphoid collections containing DC and T and B lymphocytes in the nodule suggests that local presentation of antigen may not occur in the rheumatoid nodule, as is thought to be the case in synovial membranes containing lymphoid follicles. This difference could potentially be explained by different states of activation, and differentiation of DC within the 2 lesions.
    The Journal of Rheumatology 03/2000; 27(2):339-46. · 3.26 Impact Factor
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    ABSTRACT: As only a few cell surface markers for dendritic cells (DC) have been identified to date, this study examined the expression of ligands for lectin on different human DC populations. The ability of Concanavalin A (Con A), Wheat Germ Agglutinin (WGA), peanut agglutinin (PNA), and Helix pomatia (HPA) to bind to cell lines and PBMC and DC populations was analyzed by flow cytometry and specificity of binding confirmed using inhibitory and noninhibitory sugars. The cell lines showed non-lineage-restricted binding with Con A and WGA, independent of sialidase treatment. HPA and PNA bound to a restricted number of lines, but showed broad reactivity after sialidase treatment. The peripheral blood mononuclear cells (PBMC) and directly isolated blood DC, activated CD83(+) blood DC, epidermal Langerhans cells (LC), and monocyte-derived DC (Mo-DC) showed strong binding of Con A and WGA, both before and after sialidase treatment. No HPA binding ligands were detected on PBMC populations, including directly isolated blood DC. Following sialidase treatment CD3(+), CD16(+), and a subset of CD19(+) lymphocytes bound HPA. The lectin PNA bound weakly to CD14(+) monocytes and a subpopulation of circulating DC that were HLA-DR(hi)CDw123 Dr(hi)CDw123(dim)/(neg)CD11c(+). The HLA-DR(mod)CDw123(hi)CD11c(neg) subpopulation did not bind PNA. Without sialidase treatment LC expressed both HPA and PNA ligands, but these were either absent on activated CD83(+) blood DC or weakly expressed on Mo-DC. Following sialidase treatment PBMC populations, activated CD83(+) blood DC, and Mo-DC became PNA positive. Thus human DC express several lectin ligands and PNA binding identifies a subset of blood DC. That may reflect discrete changes associated with stages of DC development or functional maturation.
    Cellular Immunology 03/2000; 200(1):36-44. · 1.74 Impact Factor

Publication Stats

3k Citations
439.78 Total Impact Points

Institutions

  • 1986–2008
    • Canterbury District Health Board
      • • Department of Haematology
      • • South Island Bone Marrow Transplant Unit (BMTU)
      • • Department of Immunology
      Christchurch, Canterbury Region, New Zealand
  • 2001
    • University of Zurich
      Zürich, Zurich, Switzerland
    • University of Queensland
      Brisbane, Queensland, Australia
  • 1999–2001
    • Mater Hospital
      Brisbane, Queensland, Australia
  • 2000
    • University of Otago
      • Department of Medicine (Dunedin)
      Dunedin, Otago, New Zealand
    • Queensland Institute of Medical Research
      Brisbane, Queensland, Australia
  • 1999–2000
    • Mater Medical Research Institute
      South Brisbane, Queensland, Australia