Medicine, Dentistry & Health Sciences Collected Works - Research Publications

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    Occult axillary node metastases in breast cancer: Their detection and prognostic significance
    McGuckin, MA ; Cummings, MC ; Walsh, MD ; Hohn, BG ; Bennett, IC ; Wright, RG (STOCKTON PRESS, 1996-01)
    Although the presence of axillary node metastases in breast cancer is a key prognostic indicator and may influence treatment decisions, a significant proportion of patients diagnosed as axillary node negative (ANN) using standard histopathological techniques may have occult nodal metastases (OMs). A combination of limited step-sectioning (4 x 100 microns intervals) and immunohistochemical staining (with cytokeratin (MNF.116) and MUC1 (BC2) antibodies) was used to detect OM in a retrospective series of 208 ANN patients. OMs were found in 53 patients (25%), and both step-sectioning and immunohistochemical detection significantly improved detection (P < 0.05). Detection using BC2 (25%) was superior to MNF.116 (18%) and haematoxylin and eosin (H&E) (8%). OMs were found in 51 patients using only the first and deepest sectioning levels and BC2 staining. OMs were more frequently found in lobular (38%) than ductal carcinoma (25%), and more frequently in women less than 50 years (41%) than in older women (19%). Univariate overall and disease-free survival analyses showed that the presence, size and number of OM had prognostic significance as did tumour size (disease-free only) and histological and nuclear grade (P > 0.05). Cox multivariate proportional hazard regression analyses showed that the presence and increasing size of OMs were significantly associated with poorer disease-free survival, independently of other prognostic factors (P < 0.05). However there was not a significant independent association of the presence of occult metastases with overall survival (P = 0.11). These findings have important implications with regard to selection of ANN patients for adjuvant therapy.
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    MONOCLONAL-ANTIBODIES REACTING WITH THE MUC2 MUCIN CORE PROTEIN
    DEVINE, PL ; MCGUCKIN, MA ; BIRRELL, GW ; WHITEHEAD, RH ; SACHDEV, GP ; SHIELD, P ; WARD, BG (STOCKTON PRESS, 1993-06)
    This study sought to produce monoclonal antibodies (MAbs) which reacted with the MUC2 core protein. Two MAbs [3A2 (IgG1) and 4F1 (IgM)] were produced by immunising female BALB/c mice with gel-formed mucin from the LS174T colon cancer cell line followed by a KLH conjugate of a 29 amino acid synthetic peptide whose sequence was derived from the variable number of tandem repeats (VNTR) region of a MUC2 cDNA clone. The MAbs reacted with synthetic MUC2 VNTR peptides but not synthetic MUC1 or MUC3 VNTR peptides, and showed specific reactivity in Western blotting with a high molecular weight protein produced by the LS174T colon carcinoma cell line. The use of shorter peptides indicated that the minimum peptide epitopes for these MAbs were different. Mab 3A2 reacted with amino acids 5-19 of the MUC2 VNTR by inhibition ELISA but not by direct ELISA, while 4F1 reacted with this peptide in both assays. Furthermore, 4F1 reacted in direct ELISA when a larger (29 amino acid) MUC2-derived peptide was coated onto the assay plate by incubating in carbonate buffer or by drying the peptide onto the assay plate, while 3A2 only reacted when this peptide was coated in carbonate buffer. The different specificity of the MAbs was also illustrated by the reactivity of 4F1 but not 3A2 with partially deglycosylated cystic fibrosis mucin. Immunohistochemical analysis with these MAbs revealed a strong reactivity with lung, gastric and colon tumours relative to normal tissue, with some breast and ovarian tumours also reacting. Both MAbs stained some normal goblet cells in the perinuclear region but not the mucin droplet or secreted mucin, indicating a reaction with immature (poorly glycosylated) mucin in the endoplasmic reticulum and/or golgi, but not with mature (fully glycosylated) mucin. In contrast, tumours showed strong diffuse cytoplasmic staining. 4F1 also showed weak apical cytoplasmic staining in some goblet cells and stained some tumours which showed no reactivity with 3A2. These antibodies should prove useful in the study of MUC2 structure and function, and in the diagnosis of some tumours.
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    CD36 GENE-TRANSFER CONFERS CAPACITY FOR PHAGOCYTOSIS OF CELLS UNDERGOING APOPTOSIS
    REN, Y ; SILVERSTEIN, RL ; ALLEN, J ; SAVILL, J (ROCKEFELLER UNIV PRESS, 1995-05-01)
    Phagocyte recognition and ingestion of intact cells undergoing apoptosis are key events in this generally important program of cell death. Insufficient phagocyte capacity for apoptotic cells can result in failure to clear dying cells before membrane integrity is lost, resulting in leakage of noxious cell contents and severe tissue damage. However, no means has been available to increase phagocytic clearance of apoptotic cells. We now report that transfection of the macrophage adhesion molecule CD36 into human Bowes melanoma cells specifically conferred greatly increased capacity to ingest apoptotic neutrophils, lymphocytes, and fibroblasts, comparable to that exhibited by macrophages. Furthermore, when CD36 was transfected into another cell type with limited capacity to take up apoptotic bodies, the monkey COS-7 cell, similar effects were observed. Therefore, CD36 gene transfer can confer "professional" capacity to ingest apoptotic cells upon "amateur" phagocytes.
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    THE SPECIFIC RECOGNITION BY MACROPHAGES OF CD8(+),CD45RO(+) T-CELLS UNDERGOING APOPTOSIS - A MECHANISM FOR T-CELL CLEARANCE DURING RESOLUTION OF VIRAL-INFECTIONS
    AKBAR, AN ; SAVILL, J ; GOMBERT, W ; BOFILL, M ; BORTHWICK, NJ ; WHITELAW, F ; GRUNDY, J ; JANOSSY, G ; SALMON, M (ROCKEFELLER UNIV PRESS, 1994-11-01)
    During viral infections, CD8+,CD45RO+ T populations expand. These primed cells express abundant levels of cytoplasmic granules that contain perforin and TIA-1. Recent work has suggested that the majority of this CD8+ population downregulates Bcl-2 protein expression and is destined to undergo apoptosis. In this study we have investigated the elimination of these apoptotic CD8+ T cells by both human monocyte-derived and murine bone marrow macrophages. We have found that these phagocytes recognize and ingest both apoptotic CD8+ and CD4+ T cells using an alpha v beta 3 (vitronectin receptor)/CD36/thrombospondin recognition system, with the same receptors being used in the recognition of apoptotic neutrophils. These data provide new evidence for a mechanism that enables the clearance of greatly increased populations of CD8+ effector cells which are found during viral infections. This enables cellular homeostasis to occur in the host upon resolution of viral diseases in vivo.
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    Measuring recent thymic emigrants in blood of normal and HIV-1-infected individuals before and after effective therapy
    Zhang, LQ ; Lewin, SR ; Markowitz, M ; Lin, HH ; Skulsky, E ; Karanicolas, R ; He, YX ; Jin, X ; Tuttleton, S ; Vesanen, M ; Spiegel, H ; Kost, R ; van Lunzen, J ; Stellbrink, HJ ; Wolinsky, S ; Borkowsky, W ; Palumbo, P ; Kostrikis, LG ; Ho, DD (ROCKEFELLER UNIV PRESS, 1999-09-06)
    The role of the thymus in HIV-1 pathogenesis remains unclear. We developed an assay to quantify the number of recent thymic emigrants in blood based on the detection of a major excisional DNA byproduct (termed alpha1 circle) of T cell receptor rearrangement. By studying 532 normal individuals, we found that alpha1 circle numbers in blood remain high for the first 10-15 yr of life, a sharp drop is seen in the late teen years, and a gradual decline occurs thereafter. Compared with age-matched uninfected control individuals, alpha1 circle numbers in HIV-1-infected adults were significantly reduced; however, there were many individuals with normal alpha1 circle numbers. In 74 individuals receiving highly active antiretroviral therapy, we found no appreciable effect on alpha1 circle numbers in those whose baseline values were already within the normal range, but significant increases were observed in those with a preexisting impairment. The increases in alpha1 circle numbers were, however, numerically insufficient to account for the rise in levels of naive T lymphocytes. Overall, it is difficult to invoke thymic regenerative failure as a generalized mechanism for CD4 lymphocyte depletion in HIV-1 infection, as alpha1 circle numbers are normal in a substantial subset of HIV-1-infected individuals.
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    Dramatic rise in plasma viremia after CD8+ T cell depletion in simian immunodeficiency virus-infected macaques
    Jin, X ; Bauer, DE ; Tuttleton, SE ; Lewin, S ; Gettie, A ; Blanchard, J ; Irwin, CE ; Safrit, JT ; Mittler, J ; Weinberger, L ; Kostrikis, LG ; Zhang, LQ ; Perelson, AS ; Ho, DD (ROCKEFELLER UNIV PRESS, 1999-03-15)
    To determine the role of CD8(+) T cells in controlling simian immunodeficiency virus (SIV) replication in vivo, we examined the effect of depleting this cell population using an anti-CD8 monoclonal antibody, OKT8F. There was on average a 99.9% reduction of CD8 cells in peripheral blood in six infected Macaca mulatta treated with OKT8F. The apparent CD8 depletion started 1 h after antibody administration, and low CD8 levels were maintained until day 8. An increase in plasma viremia of one to three orders of magnitude was observed in five of the six macaques. The injection of a control antibody to an infected macaque did not induce a sustained viral load increase, nor did it significantly reduce the number of CD8(+) T cells. These results demonstrate that CD8 cells play a crucial role in suppressing SIV replication in vivo.