87 results on '"Grundy JE"'
Search Results
2. Bone marrow transplant recipients have defective MHC-unrestricted cytotoxic responses against cytomegalovirus in comparison with Epstein- Barr virus: the importance of target cell expression of lymphocyte function-associated antigen 1 (LFA1)
- Author
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Duncombe, AS, primary, Grundy, JE, additional, Oblakowski, P, additional, Prentice, HG, additional, Gottlieb, DJ, additional, Roy, DM, additional, Reittie, JE, additional, Bello-Fernandez, C, additional, Hoffbrand, AV, additional, and Brenner, MK, additional
- Published
- 1992
- Full Text
- View/download PDF
3. Gamma-interferon and tumor necrosis factor production after bone marrow transplantation is augmented by exposure to marrow fibroblasts infected with cytomegalovirus
- Author
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Duncombe, AS, primary, Meager, A, additional, Prentice, HG, additional, Grundy, JE, additional, Heslop, HE, additional, Hoffbrand, AV, additional, and Brenner, MK, additional
- Published
- 1990
- Full Text
- View/download PDF
4. Evaluation of the benchmark dose for point of departure determination for a variety of chemical classes in applied regulatory settings.
- Author
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Izadi H, Grundy JE, and Bose R
- Subjects
- Canada, Humans, No-Observed-Adverse-Effect Level, Retrospective Studies, Risk Assessment, Benchmarking
- Abstract
Repeated-dose studies received by the New Substances Assessment and Control Bureau (NSACB) of Health Canada are used to provide hazard information toward risk calculation. These studies provide a point of departure (POD), traditionally the NOAEL or LOAEL, which is used to extrapolate the quantity of substance above which adverse effects can be expected in humans. This project explored the use of benchmark dose (BMD) modeling as an alternative to this approach for studies with few dose groups. Continuous data from oral repeated-dose studies for chemicals previously assessed by NSACB were reanalyzed using U.S. EPA benchmark dose software (BMDS) to determine the BMD and BMD 95% lower confidence limit (BMDL(05) ) for each endpoint critical to NOAEL or LOAEL determination for each chemical. Endpoint-specific benchmark dose-response levels , indicative of adversity, were consistently applied. An overall BMD and BMDL(05) were calculated for each chemical using the geometric mean. The POD obtained from benchmark analysis was then compared with the traditional toxicity thresholds originally used for risk assessment. The BMD and BMDL(05) generally were higher than the NOAEL, but lower than the LOAEL. BMDL(05) was generally constant at 57% of the BMD. Benchmark provided a clear advantage in health risk assessment when a LOAEL was the only POD identified, or when dose groups were widely distributed. Although the benchmark method cannot always be applied, in the selected studies with few dose groups it provided a more accurate estimate of the real no-adverse-effect level of a substance., (© 2011 Society for Risk Analysis.)
- Published
- 2012
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5. Plasminogen binds to plasmin-modulated factor Xa by Ca(2+) - and C-terminal lysine-dependent and -independent interactions.
- Author
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Grundy JE, Hancock MA, Meixner SC, MacKenzie RC, Koschinsky ML, and Pryzdial EL
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- Calcium pharmacology, Fibrinolysin, Humans, Pancreatic Elastase metabolism, Peptide Fragments chemistry, Protein Binding, Surface Plasmon Resonance, Calcium metabolism, Factor Xa metabolism, Lysine, Peptide Fragments metabolism, Plasminogen metabolism
- Abstract
Plasminogen binding to receptors involves both C-terminal lysine- dependent and -independent interactions. The latter are poorly understood. Our earlier work demonstrated a novel Ca (2+) -enhanced bivalent interaction between plasmin-cleaved FXa (FXa33/13) and plasminogen truncated at Lys78 (Lys-Pg). Here we hypothesized that the effects of Ca (2+) may enable dissection of the C-terminal lysine-dependent and -independent interactions. To evaluate the role of the Glu-plasminogen (Glu-Pg) amino acids 1 - 77, binding of FXa33/13 to immobilized Glu-Pg was compared to Lys-Pg by surface plasmon resonance. Under identical conditions, approximately half the amount of FXa33/13 bound to Glu-Pg. The simplest fit of data suggested a 2:1 plasminogen:FXa33/13 stoichiometry for both, which were proportionately enhanced by Ca (2+) . Only Lys-Pg demonstrated significant Ca (2+) -independent binding to FXa33/13. In the presence of Ca (2+) , weak C-terminal lysine-independent binding could be detected, but only for Glu-Pg. The elastase-generated plasminogen fragment encompassing the angiostatin-like kringle domains 1 to 3 (K1 - 3) inhibited binding of FXa33/13 to Lys-Pg, whereas fragments corresponding to kringle 4- and kringle 5-protease domain had no effect. Immobilized K1 - 3 binding to FXa33/13 had both Ca (2+) -dependent and -independent components. The principal K (d) for the interaction was 10-fold higher than Lys-Pg. In the presence of Ca (2+) , eACA inhibited FXa33/13 binding to K1 - 3 by 30%, but eliminated binding in the absence of Ca (2+) . These studies suggest that Ca (2+) -dependent and -independent binding of Lys-Pg to FXa33/13 are C-terminal lysine-dependent. The N-terminal 1 - 77 amino acids of Glu-Pg confer significant C-terminal lysine-independent binding, which may play a role during the initiating stages of plasminogen activation.
- Published
- 2007
6. Coagulation factor Va Glu-96-Asp-111: a chelator-sensitive site involved in function and subunit association.
- Author
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Zeibdawi AR, Grundy JE, Lasia B, and Pryzdial EL
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- Amino Acid Sequence, Aspartic Acid genetics, Binding Sites, Chelating Agents pharmacology, Dimerization, Factor Va metabolism, Glutamic Acid genetics, Humans, Molecular Sequence Data, Mutagenesis, Site-Directed, Phospholipids metabolism, Protein Subunits, Thromboplastin metabolism, Aspartic Acid physiology, Factor Va chemistry, Factor Va physiology, Glutamic Acid physiology
- Abstract
Coagulation FVa (factor Va) accelerates the essential generation of thrombin by FXa (factor Xa). Although the noncovalent Ca2+-dependent association between the FVa light and heavy subunits (FVaL and FVaH) is required for function, little is known about the specific residues involved. Previous fragmentation studies and homology modelling led us to investigate the contribution of Leu-94-Asp-112. Including prospective divalent cation-binding acidic amino acids, nine conserved residues were individually replaced with Ala in the recombinant B-domainless FVa precursor (DeltaFV). While mutation of Thr-104, Glu-108, Asp-112 or Tyr-100 resulted in only minor changes to FXa-mediated thrombin generation, the functions of E96A (81%), D111A (70%) and D102A (60%) mutants (where the single-letter amino acid code is used) were notably reduced. The mutants targeting neighbouring acidic residues, Asp-79 and Glu-119, had activity comparable with DeltaFV, supporting the specific involvement of select residues. Providing a basis for reduced activity, thrombin treatment of D111A resulted in spontaneous dissociation of subunits. Since FVaH and FVaL derived from E96A or D102A remained associated in the presence of Ca2+, like the wild type, but conversely dissociated rapidly upon chelation, a subtle difference in divalent cation co-ordination is implied. Subunit interactions for all other single-point mutants resembled the wild type. These data, along with corroborating multipoint mutants, reveal Asp-111 as essential for FVa subunit association. Although Glu-96 and Asp-102 can be mutated without gross changes to divalent cation-dependent FVaH-FVaL interactions, they too are required for optimal function. Thus Glu-96-Asp-111 imparts at least two discernible effects on FVa coagulation activity.
- Published
- 2004
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7. Ca2+-dependent and phospholipid-independent binding of annexin 2 and annexin 5.
- Author
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Brooks ND, Grundy JE, Lavigne N, Derry MC, Restall CM, MacKenzie CR, Waisman DM, and Pryzdial EL
- Subjects
- Protein Binding, Surface Plasmon Resonance, Annexin A2 metabolism, Annexin A5 metabolism, Calcium metabolism, Phospholipids metabolism
- Abstract
Annexins are a family of homologous proteins that associate with anionic phospholipid (aPL) in the presence of Ca(2+). Evidence that the function of one annexin type may be regulated by another was recently reported in studies investigating cytomegalovirus-aPL interactions, where the fusogenic function of annexin 2 (A2) was attenuated by annexin 5 (A5). This observation suggested that A2 may bind directly to A5. In the present study, we demonstrated this interaction. The A2-A5 complex was first detected utilizing (covalently linked) fluorescein-labelled A5 (F-A5) as a reporter group. The interaction required concentrations of Ca(2+) in the millimolar range, had an apparent dissociation constant [ K (d)(app)] of 1 nM at 2 mM Ca(2+) and was independent of aPL. A2 bound comparably with F-A5 pre-equilibrated with an amount of aPL that could bind just the F-A5 or to an excess amount of aPL providing sufficient binding sites for all of F-A5 and A2. A2-A5 complex formation was corroborated in an experiment, where [(125)I]A2 associated in a Ca(2+)-dependent manner with A5 coated on to polystyrene. Surface plasmon resonance was used as a third independent method to demonstrate the binding of A2 and A5 and, furthermore, supported the conclusion that the monomeric and tetrameric forms of A2 bind equivalently to A5. Together these results demonstrate an A2-A5 interaction and provide an explanation as to how A5 inhibits the previously reported A2-dependent enhancement of virus-aPL fusion.
- Published
- 2002
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8. Binding of plasminogen and tissue plasminogen activator to plasmin-modulated factor X and factor Xa.
- Author
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Grundy JE, Lavigne N, Hirama T, MacKenzie CR, and Pryzdial EL
- Subjects
- Anions, Binding Sites, Calcium metabolism, Dimerization, Humans, Hydrolysis, Kinetics, Peptide Fragments metabolism, Phospholipids metabolism, Protein Binding, Surface Plasmon Resonance, Factor X metabolism, Factor Xa metabolism, Fibrinolysin metabolism, Plasminogen metabolism, Tissue Plasminogen Activator metabolism
- Abstract
Previous work in our laboratory has suggested that the fibrinolytic enzyme plasmin (Pn) inactivates coagulation factors X (FX) and Xa (FXa) in the presence of Ca(2+) and anionic phospholipid (aPL), producing fragments which bind plasminogen (Pg) and accelerate tissue plasminogen activator (t-PA). Our goals here were to determine if the Pn-mediated fragments of FX or FXa remain associated, whether they directly bind t-PA, and to quantify their interaction with Pg. Binding to aPL, benzamidine-Sepharose, or the active-site inhibitor dansyl-Glu-Gly-Arg-chloromethyl ketone demonstrated that Pn cleavage yielded noncovalent heterodimers of a fragment containing the aPL-binding domain (FXgamma(47) or FXagamma(33)) and a 13-kDa fragment (FXgamma(13) or FXagamma(13)). Both ligand blotting and surface plasmon resonance (SPR) showed that Pn-cleaved FX and FXa bound t-PA directly when Pn-treatment was effected in the presence of aPL and Ca(2+). Using SPR, apparent K(d) values of 1-3 microM and 0.3-0.4 microM were measured directly and by competition for the FXgamma(47/13)-Pg and FXagamma(33/13)-Pg interactions, respectively. For the first time, Pg-binding to a receptor was shown to be Ca(2+) enhanced, although primarily mediated by C-terminal lysine residues. Mathematical modeling of kinetic data suggesting two Pg per FXgamma(47/13) or FXagamma(33/13) was consistent with our conclusion that each subunit of FXgamma(47/13) or FXagamma(33/13) contains a C-terminal lysine. Earlier X-ray structures show that these Lys residues are distal from each other and the membrane, supporting the model where each interacts with a separate Pg. t-PA acceleration by FXgamma(47/13) or FXagamma(33/13) may therefore involve simultaneous presentation of two substrate molecules.
- Published
- 2001
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9. Identification of three HLA-A*0201-restricted cytotoxic T cell epitopes in the cytomegalovirus protein pp65 that are conserved between eight strains of the virus.
- Author
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Solache A, Morgan CL, Dodi AI, Morte C, Scott I, Baboonian C, Zal B, Goldman J, Grundy JE, and Madrigal JA
- Subjects
- Antigen Presentation, Cell Line, Cell Line, Transformed, Cytomegalovirus isolation & purification, Epitopes, T-Lymphocyte chemistry, HLA-A2 Antigen chemistry, Humans, Lymphocyte Activation, Peptide Fragments chemistry, Peptide Fragments metabolism, Phosphoproteins chemistry, Phosphoproteins immunology, Protein Binding immunology, Species Specificity, T-Lymphocytes, Cytotoxic immunology, Viral Matrix Proteins chemistry, Viral Matrix Proteins immunology, Conserved Sequence immunology, Cytomegalovirus immunology, Epitopes, T-Lymphocyte metabolism, HLA-A2 Antigen metabolism, Phosphoproteins metabolism, T-Lymphocytes, Cytotoxic metabolism, Viral Matrix Proteins metabolism
- Abstract
The Ag specificity of the CTL response against CMV is directed almost entirely to a single CMV tegument protein, the phosphoprotein pp65. We report the identification of three peptides derived from the protein pp65 that displayed a high or intermediate binding to HLA-A*0201 molecules, which were also able to induce an in vitro CTL response in peripheral blood lymphocytes from CMV seropositive individuals. The peptide-specific CTLs generated were capable of recognizing the naturally processed pp65 either presented by CMV-infected cells or by cells infected with an adenovirus construct expressing pp65 in an HLA-A*0201-restricted manner. Thus, we were able to demonstrate responses to subdominant CTL epitopes in CMV-pp65 that were not detected in polyclonal cultures obtained by conventional stimulations. We also found that the amino acid sequences of the three peptides identified as HLA-A*0201-restricted CTL epitopes were conserved among different wild-type strains of CMV obtained from renal transplant patients, an AIDS patient, and a congenitally infected infant, as well as three laboratory strains of the virus (AD169, Towne and Davis). These observations suggest that these pp65 CTL peptide epitopes could potentially be used as synthetic peptide vaccines or for other therapeutic strategies aimed at HLA-A*0201-positive individuals, who represent approximately 40% of the European Caucasoid population. However, strain variation must be taken in consideration when the search for CTL epitopes is extended to other HLA class I alleles, because these mutations may span potential CTL epitopes for other HLA molecules, as it is described in this study.
- Published
- 1999
10. Two clinical isolates and the Toledo strain of cytomegalovirus contain endothelial cell tropic variants that are not present in the AD169, Towne, or Davis strains.
- Author
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MacCormac LP and Grundy JE
- Subjects
- Cells, Cultured, Cytomegalovirus growth & development, Cytomegalovirus isolation & purification, Endothelium, Vascular cytology, Fibroblasts cytology, Fibroblasts virology, Humans, Cytomegalovirus physiology, Genetic Variation
- Abstract
The highly fibroblast-passaged AD169, Towne, and Davis strains of cytomegalovirus (CMV) were found to have a restricted capacity to infect endothelial cells in vitro. Although such replication could be increased by a combination of low speed centrifugation and sodium butyrate treatment, the extracellular virus produced was infectious for fibroblasts but not for endothelial cells. In contrast, the low passage Toledo strain, and a low passage fibroblast-grown clinical isolate of CMV, C1F, could be continually passaged in endothelial cells, giving the strains C1FE and Toledo.E. Whilst, using the conditions described above, initial infection of endothelial cells with AD169 or C1F resulted in similar titres of extracellular virus as assayed on fibroblasts, only the virus from the C1F strain was infectious for endothelial cells. Passage of C1F in fibroblasts decreased its ability to infect endothelial cells, whilst retaining equal ability to infect fibroblasts. Although endothelial-cell-passaged cell-free C1FE virus was endothelial cell-tropic, it was still much more infectious for fibroblasts than for endothelial cells. It is concluded that the C1F and Toledo strains, but not the AD169, Towne, or Davis strains, contained endothelial cell tropic variants, which could be lost on passage through fibroblasts, but retained on passage through endothelial cells. Furthermore, virus in an ex vivo source of CMV, a blood specimen, was found to be more tropic for fibroblasts than for endothelial cells, suggesting that in vivo CMV exists as quasi strains with different cell tropism, some of which might be lost in vitro by passage in an inappropriate cell type.
- Published
- 1999
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11. Natural killer cell lysis of cytomegalovirus (CMV)-infected cells correlates with virally induced changes in cell surface lymphocyte function-associated antigen-3 (LFA-3) expression and not with the CMV-induced down-regulation of cell surface class I HLA.
- Author
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Fletcher JM, Prentice HG, and Grundy JE
- Subjects
- Antibodies, Blocking pharmacology, Antibodies, Monoclonal pharmacology, CD2 Antigens immunology, CD58 Antigens drug effects, CD58 Antigens metabolism, Cell Line, Cytomegalovirus classification, Cytomegalovirus drug effects, Cytotoxicity Tests, Immunologic, Disease Susceptibility, Endothelium, Vascular, Fibroblasts drug effects, Fibroblasts immunology, Fibroblasts virology, Ganciclovir pharmacology, HLA Antigens biosynthesis, Histocompatibility Antigens Class I biosynthesis, Humans, Immunity, Cellular, Immunity, Innate drug effects, Kinetics, Lymphocyte Activation, Tumor Cells, Cultured, CD58 Antigens biosynthesis, Cytomegalovirus immunology, Cytotoxicity, Immunologic drug effects, Down-Regulation immunology, HLA Antigens metabolism, Histocompatibility Antigens Class I metabolism, Killer Cells, Natural immunology
- Abstract
CMV and other viruses down-regulate the cell surface expression of class I HLA, and while this allows them to evade CTL, it may make infected cells more susceptible to lysis by NK cells, due to the failure to engage class I inhibitory receptors on the NK cell. We studied CMV infection and found that fibroblasts infected with virus strains Towne, Toledo, Davis, and C1FE were refractory to NK lysis, while those infected with strains AD169, C1F, or R7 were susceptible. All viral strains down-regulated class I HLA to a similar extent, and we concluded that there was no evidence for any correlation between the latter and susceptibility to NK lysis. In contrast, there was a strong correlation between NK killing of CMV-infected cells and cell surface levels of lymphocyte function-associated antigen-3 (LFA-3). Fibroblasts infected with the Towne, Toledo, Davis, and C1FE strains of CMV down-regulated LFA-3 expression and were refractory to lysis, while strains AD169, C1F, and R7 up-regulated LFA-3 and were susceptible to NK killing. U373 MG (malignant glioma) cells expressed constitutively high levels of LFA-3 and were sensitive to NK lysis when infected with any of the above-listed CMV strains. We estimated that a minimum of between 29,000 and 71,000 LFA-3 molecules per target cell were needed for NK susceptibility. The effects on LFA-3 expression were due to immediate early/early viral gene products. We also demonstrated that fibroblasts infected with the strains Towne, Toledo, Davis, and C1FE expressed a ganciclovir-sensitive late CMV gene product, which delivered an inhibitory signal to NK cells.
- Published
- 1998
12. Cytomegalovirus-infected endothelial cells recruit neutrophils by the secretion of C-X-C chemokines and transmit virus by direct neutrophil-endothelial cell contact and during neutrophil transendothelial migration.
- Author
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Grundy JE, Lawson KM, MacCormac LP, Fletcher JM, and Yong KL
- Subjects
- Cell Communication, Cell Movement, Cells, Cultured, Chemokine CXCL1, Chemokines, CXC immunology, Chemotactic Factors biosynthesis, Chemotactic Factors immunology, Chemotaxis, Leukocyte, Coculture Techniques, Culture Media, Conditioned, Cytomegalovirus immunology, Endothelium, Vascular metabolism, Growth Substances biosynthesis, Growth Substances immunology, Humans, Interleukin-8 biosynthesis, Phosphoproteins metabolism, Viral Matrix Proteins metabolism, Chemokines, CXC metabolism, Cytomegalovirus physiology, Endothelium, Vascular immunology, Endothelium, Vascular virology, Intercellular Signaling Peptides and Proteins, Neutrophils immunology, Neutrophils virology
- Abstract
Infection of endothelial cells with an endothelial cell-tropic clinical isolate of cytomegalovirus (CMV), C1FE, induced enhanced production of the neutrophil chemoattractant C-X-C chemokines interleukin-8 and GROalpha. Infected endothelial cell supernatants induced neutrophil chemotaxis in a transendothelial migration assay. Neutrophils acquired the CMV structural protein pp65 following either coculture with infected endothelial cells or transmigration through infected endothelium. The lack of CMV p72 expression in the neutrophils indicated that viral replication had not occurred in these cells. Of importance, neutrophils acquired infectious CMV during transmigration across infected endothelium and were subsequently able to transmit infectious virus to fibroblasts. Thus, CMV-infected endothelial cells can recruit neutrophils by the secretion of C-X-C chemokines and can transmit the virus to them by direct cell-to-cell contact and during neutrophil transendothelial migration, suggesting that the neutrophil-endothelial cell interaction plays an important role in virus dissemination in vivo.
- Published
- 1998
- Full Text
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13. Addition of a poly-(6X) His tag to Milk Bundle-1 and purification using immobilized metal-affinity chromatography.
- Author
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Grundy JE, Wirtanen LY, and Beauregard M
- Subjects
- Amino Acid Sequence, Base Sequence, Cloning, Molecular, Dietary Proteins analysis, Dietary Proteins genetics, Electrophoresis, Polyacrylamide Gel, Escherichia coli genetics, Histidine chemistry, Molecular Sequence Data, Oligodeoxyribonucleotides, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins isolation & purification, Chromatography, Affinity methods, Dietary Proteins isolation & purification, Peptides chemistry
- Abstract
Milk Bundle 1 (MB-1) is a de novo designed protein enriched in M, T, K, and L. Its future application is as a high-quality dietary protein source for ruminants. The protein is currently expressed in Escherichia coli and is being characterized to solve its folded conformation. MB-1 has marginal stability at room temperature, which has hindered our attempts at characterization. To increase the stability of the protein at room temperature, the purification procedure was examined and changed to hopefully increase its effectiveness. We describe here the production and purification of a new MB-1 with six His residues at the C-terminal end. This allows the new mutant (MB-1-His) to bind metal ions and to be purified with immobilized metal-affinity chromatography (IMAC). MB-1-His obtained using IMAC was purer on SDS-PAGE than both MB-1 or MB-1-His isolated using the current protocol. The IMAC protocol is more economical and more efficient; preliminary results show that the protein purified by this method is also quite stable at room temperature.
- Published
- 1998
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14. Antioxidant defenses and lipid peroxidation damage in estivating toads, Scaphiopus couchii.
- Author
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Grundy JE and Storey KB
- Subjects
- Animals, Antioxidants analysis, Body Water physiology, Glutathione analysis, Oxygen Consumption physiology, Reactive Oxygen Species metabolism, Xanthine Dehydrogenase metabolism, Xanthine Oxidase metabolism, Anura physiology, Enzymes metabolism, Estivation physiology, Lipid Peroxidation physiology
- Abstract
Tissue-specific changes in antioxidant defenses and lipid peroxidation damage were analyzed in spadefoot toads, Scaphiopus couchii, to determine how these responded during estivation, a state of suppressed oxygen consumption. Maximal activities of glutathione-S-transferase, glutathione reductase, glutathione peroxidase, superoxide dismutase and catalase were measured in six organs from 2-month-estivated toads and compared with activities in animals awakened for 10 days after estivation. Activities of many enzymes, particularly the glutathione-linked enzymes, were significantly lower in tissues of estivating toads than in awake toads. This indicates that enzymatic antioxidant defenses are probably modulated in response to the rate of reactive oxygen species generation in tissues, which is proportional to oxygen consumption. Antioxidant enzyme activities were largely insensitive to high urea, which accumulates during estivation, but were inhibited by elevated KCl. Levels of reduced glutathione were also significantly lower in three organs during estivation and all organs, except skeletal muscle, exhibited a higher oxidized/reduced glutathione ratio, indicating a more oxidized state during estivation. Products of lipid peroxidation (conjugated dienes, lipid hydroperoxides) were higher in tissues of estivated than control toads, suggesting accumulated oxidative damage to lipids during dormancy. One enzymatic source of free radical generation, xanthine oxidase, appeared to have little impact because its activity was detectable only in liver and was significantly lower in estivated toads. The data indicate that both enzymatic and metabolite antioxidant defenses in toads are adaptable systems that are modulated in estivating versus awake states.
- Published
- 1998
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15. Human cytomegalovirus infection up-regulates interleukin-8 gene expression and stimulates neutrophil transendothelial migration.
- Author
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Craigen JL, Yong KL, Jordan NJ, MacCormac LP, Westwick J, Akbar AN, and Grundy JE
- Subjects
- Blotting, Northern, Cell Adhesion Molecules metabolism, Cell Culture Techniques, Endothelium, Vascular immunology, Fibroblasts immunology, Humans, Interleukin-8 genetics, RNA, Messenger genetics, Chemotaxis, Leukocyte immunology, Cytomegalovirus Infections immunology, Interleukin-8 biosynthesis, Neutrophils immunology, Up-Regulation immunology
- Abstract
Virus-induced alterations in the cellular expression of chemokines may be important in directing the migration of specific leucocyte subsets to sites of infection, thereby playing a pivotal role in viral pathogenesis. We show here that cytomegalovirus (CMV) infection of human fibroblasts resulted in significantly increased expression of the C-X-C or alpha-chemokine interleukin-8 (IL-8), at both the mRNA and protein levels. Increased IL-8 production was seen following infection with the high passage laboratory CMV strains AD169, Towne, or Davis, as well as the low passage clinical CMV isolates Toledo or C1F. The increase in IL-8 production had functional consequences, as demonstrated by the ability of supernatants from CMV-infected fibroblasts to significantly enhance neutrophil transendothelial migration. The latter was independent of alterations in adhesion molecule expression on the endothelial cells, and was abrogated by neutralizing antibodies specific for IL-8. Direct infection of endothelium with the endothelial cell-tropic CMV strain C1FE, also resulted in enhanced neutrophil transendothelial migration. Neutrophils play an important role in the dissemination of CMV throughout the body, and thus CMV-induced neutrophil recruitment would be expected to enhance CMV dissemination. Increased production of chemokines in response to CMV infection could also disrupt the fine balance between a beneficial and a destructive immune response, thereby potentially contributing to pathology.
- Published
- 1997
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16. A humanized antibody against human cytomegalovirus (CMV) gpUL75 (gH) for prophylaxis or treatment of CMV infections.
- Author
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Hamilton AA, Manuel DM, Grundy JE, Turner AJ, King SI, Adair JR, White P, Carr FJ, and Harris WJ
- Subjects
- Amino Acid Sequence, Animals, Antibodies, Viral chemistry, Base Sequence, Cell Line, Cytomegalovirus Infections drug therapy, Humans, Immunoglobulin Variable Region chemistry, Mice, Molecular Sequence Data, Recombinant Fusion Proteins immunology, Antibodies, Viral immunology, Cytomegalovirus Infections prevention & control, Viral Envelope Proteins immunology
- Abstract
A humanized monoclonal antibody that binds to the 86-kDa glycoprotein, gpUL75 (gH), of human cytomegalovirus (CMV) has been developed. The six complementarity determining regions of the heavy and light chains of the mouse antibody HCMV16 were transferred to human antibody framework sequences and combined with human antibody constant regions to produce a complete antibody. The reshaped antibody recognized cells infected with a variety of virus strains and neutralized clinical isolates of CMV as efficiently as laboratory strains in a conventional plaque reduction assay. This antibody provides a potential agent for the prevention or treatment of CMV infections in humans.
- Published
- 1997
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17. Selective migration of highly differentiated primed T cells, defined by low expression of CD45RB, across human umbilical vein endothelial cells: effects of viral infection on transmigration.
- Author
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Borthwick NJ, Akbar AN, MacCormac LP, Lowdell M, Craigen JL, Hassan I, Grundy JE, Salmon M, and Yong KL
- Subjects
- Cell Line, Cell Movement immunology, Chemokines immunology, Cytokines immunology, Cytomegalovirus Infections immunology, Herpesviridae Infections immunology, Herpesvirus 4, Human, Humans, Immunophenotyping, Tumor Virus Infections immunology, Umbilical Veins immunology, Endothelium, Vascular immunology, Leukocyte Common Antigens analysis, T-Lymphocyte Subsets immunology, Virus Diseases immunology
- Abstract
Low expression of CD45RB on CD45RO+ T lymphocytes defines a subset of highly differentiated T lymphocytes that accumulate in vivo within the affected joints of patients with rheumatoid arthritis (RA). Although it is known that CD45RO+ T lymphocytes migrate to sites of inflammation in vivo, it is not clear whether within this subset the CD45RBlo cells are selectively recruited or develop in situ within the joint. Using a transwell system we show that a small proportion of resting T lymphocytes migrated across unactivated human umbilical vein endothelial cells (HUVEC). These migrating cells were CD45RO+ and enriched for low CD45RB expression. In addition, both the CD45RO+CD45RBlo subset and migrating cells expressed increased levels of beta 1 and beta 2 integrins and CD44. The percentage of CD45RO+CD45RBlo T lymphocytes was increased in the circulation of patients with acute Epstein-Barr virus (EBV) infection. These in vivo activated cells also expressed increased levels beta 1 and beta 2 integrins and CD44, and showed an enhanced rate of transmigration compared with resting T lymphocytes. Transmigration of T lymphocytes was increased using the chemokines RANTES and lymphotactin and the cytokine interleukin-15 (IL-15). In addition, infection of the HUVEC with cytomegalovirus (CMV) led to an enhanced movement of T lymphocytes. In all of these cases the selective migration of the CD45RBlo subset was maintained. Thus although the rate of T-lymphocyte transmigration could be influenced by a number factors, the CD45RO+CD45RBlo subset has a migratory advantage suggesting that more differentiated CD45RO+CD45RBlo T lymphocytes are selectively recruited to sites of inflammation.
- Published
- 1997
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18. Human cytomegalovirus induces an Fc gamma receptor (Fc gammaR) in endothelial cells and fibroblasts that is distinct from the human cellular Fc gammaRs.
- Author
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MacCormac LP and Grundy JE
- Subjects
- Animals, Antibodies, Monoclonal immunology, Butyrates pharmacology, Butyric Acid, Cells, Cultured, Endothelium cytology, Fibroblasts virology, Flow Cytometry, Fluorescent Antibody Technique, Indirect, Humans, Immunoglobulin Fc Fragments pharmacology, Immunoglobulin G immunology, Lung cytology, Membrane Proteins immunology, Mice, Nuclear Envelope immunology, Nuclear Envelope virology, Rabbits, Receptors, Cell Surface biosynthesis, Receptors, Cell Surface immunology, Receptors, IgG immunology, Trypsin pharmacology, Umbilical Veins cytology, Viral Proteins immunology, Cytomegalovirus Infections immunology, Membrane Proteins biosynthesis, Receptors, IgG analysis, Receptors, IgG biosynthesis, Viral Proteins biosynthesis
- Abstract
The expression of a trypsin-sensitive receptor for the Fc portion of IgG (Fc gammaR) was demonstrated by flow cytometry on the surface of human umbilical vein endothelial cells and fibroblasts infected with human cytomegalovirus (CMV). Double-labeling experiments showed strong expression of the CMV Fc gammaR in a perinuclear region of infected cells but not in bystander uninfected cells. The CMV Fc gammaR did not react with a panel of murine monoclonal antibodies directed against the known human IgG Fc receptors, Fc gammaRI, Fc gammaRII, and Fc gammaRIII. The cytoplasmic form but not the cell surface form of CMV Fc gammaR bound murine IgG3 moderately and murine IgG1 more weakly, while both forms bound rabbit IgG almost as strongly as human IgG. The function of CMV Fc gammaR is unclear, but it may allow CMV to evade host antibody responses. However, the binding of immune complexes to infected endothelium might also contribute to immunopathology.
- Published
- 1996
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19. Cytomegalovirus induced up-regulation of LFA-3 (CD58) and ICAM-1 (CD54) is a direct viral effect that is not prevented by ganciclovir or foscarnet treatment.
- Author
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Craigen JL and Grundy JE
- Subjects
- Antiviral Agents pharmacology, Bone Marrow Transplantation immunology, Cytomegalovirus chemistry, Cytomegalovirus Infections complications, Fibroblasts virology, Graft Rejection prevention & control, Humans, Viral Proteins biosynthesis, CD58 Antigens physiology, Cytomegalovirus physiology, Foscarnet therapeutic use, Ganciclovir therapeutic use, Intercellular Adhesion Molecule-1 physiology, Organ Transplantation physiology, Up-Regulation physiology
- Abstract
Cytomegalovirus (CMV) is a major pathogen in transplant recipients and AIDS patients, and the virus may also play a role in allograft rejection. Previous work from this laboratory demonstrated increased cell surface expression of the adhesion molecules ICAM-1 (CD54) and LFA-3 (CD58) following CMV infection in vitro. We investigated whether the induction of adhesion molecules by CMV was a direct viral effect or secondary to cytokine induction. Cytokines known to up-regulate ICAM-1, such as TNFalpha or IL-1beta, were not detected in the supernatants of infected fibroblasts, and neutralizing antibodies against these cytokines did not abrogate the induction of either ICAM-1 or LFA-3 by CMV. Infected cell supernatants had increased levels of IL-6, IL-8 and IFNbeta however, the addition of recombinant forms of these cytokines did not affect adhesion molecule expression. Neither virus-free infected cell supernatants nor UV-inactivated virus up-regulated adhesion molecules, demonstrating that the induction of ICAM-1 and LFA-3 by CMV was a direct effect requiring infectious virus. Effective antiviral treatment with ganciclovir or foscarnet accentuated rather than abrogated the up-regulation of adhesion molecules, suggesting that CMV immediate early/early gene expression, which is not blocked by such treatment, was responsible for the adhesion molecule induction. Thus, despite effective antiviral therapy in the transplant recipient, CMV infected cells may continue to provide a focus of proinflammatory activity, which could contribute to immunopathology and/or accentuate graft rejection or graft-versus-host disease in vivo.
- Published
- 1996
- Full Text
- View/download PDF
20. A three-center European external quality control study of PCR for detection of cytomegalovirus DNA in blood.
- Author
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Grundy JE, Ehrnst A, Einsele H, Emery VC, Hebart H, Prentice HG, and Ljungman P
- Subjects
- Bone Marrow Transplantation adverse effects, Cytomegalovirus genetics, Cytomegalovirus Infections diagnosis, Cytomegalovirus Infections etiology, DNA, Viral genetics, DNA, Viral standards, Diagnostic Errors, Europe, Humans, Leukocytes virology, Polymerase Chain Reaction methods, Quality Control, Viremia diagnosis, Viremia etiology, Virology methods, Virology standards, Cytomegalovirus isolation & purification, DNA, Viral blood, Polymerase Chain Reaction standards
- Abstract
The presence of cytomegalovirus (CMV) in the blood has important consequences for patient management, and an external quality control study of its detection by the PCR was conducted by the Infectious Disease Working Party of the European Group for Blood and Marrow Transplantation. Forty-eight coded peripheral blood samples from bone marrow transplant recipients were processed in parallel in three European centers by using the routine in-house PCR assay. Protocols varied in choice of primers, specificity and amplificability controls, and sample processing. Results for 38 of 47 samples agreed, 35 being negative and 3 positive. Of the 12 samples reported as positive by a least one center, only 3 were found to be positive by all three centers, 1 was found to be positive by two centers, and the remaining 8 were found to be positive by one center only. The nine discrepant samples appeared to contain around 1,000-fold less viral DNA than the three concordant positive samples. CMV detection was affected both by the number of leukocytes from which DNA was extracted and by the number of cell equivalents added per PCR. External quality control schemes for CMV PCR are clearly necessary in order to compare data from different centers, and recommendation for standardizing the PCR detection of CMV in blood leukocytes are made.
- Published
- 1996
- Full Text
- View/download PDF
21. Biochemistry below 0 degrees C: nature's frozen vertebrates.
- Author
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Storey KB, Mosser DD, Douglas DN, Grundy JE, and Storey JM
- Subjects
- Adenosine Triphosphate metabolism, Animals, Body Temperature physiology, Phosphorylases metabolism, Ranidae metabolism, Turtles metabolism, Cryopreservation, Extracellular Space physiology, Freezing, Liver ultrastructure, Magnetic Resonance Imaging
- Abstract
Although alien to man, the ability to endure the freezing of extracellular body fluids during the winter has developed in several species of terrestrially hibernating frogs and turtles as well as in many species of insects and other invertebrates. Wood frogs, for example, can endure freezing for at least 2 weeks with no breathing, no heart beat or blood circulation, and with up to 65% of their total body water as ice. Our studies are providing a comprehensive view of the requirements for natural freezing survival and of the physical and metabolic protection that must be offered for effective cryopreservation of vertebrate organs. Molecular mechanisms of natural freeze tolerance in lower vertebrates include: 1) control over ice crystal growth in plasma by ice nucleating proteins, 2) the accumulation of low molecular weight cryoprotectants to minimize intracellular dehydration and stabilize macromolecular components, and 3) good ischemia tolerance by all organs that may include metabolic arrest mechanisms to reduce organ energy requirements while frozen. Cryomicroscopy of tissue slices and magnetic resonance imaging (MRI) of whole animals is revealing the natural mode of ice propagation through an organism. MRI has also revealed that thawing is non-uniform; core organs (with high cryoprotectant levels) melt first, facilitating the early resumption of heart beat and blood circulation. Studies of the production and actions of the natural cryoprotectant, glucose, in frogs have shown its importance in maintaining a critical minimum cell volume in frozen organs and new work on the metabolic effects of whole body dehydration in 3 species of frogs has indicated that adaptations supporting freeze tolerance grew out of mechanisms that deal with desiccation resistance in amphibians. Studies of the regulation of cryoprotectant glucose synthesis by wood frog liver have shown the role of protein kinases and of alpha and beta adrenergic receptors in regulating the glycemic response, and of changes in membrane glucose transporter proteins to facilitate cryoprotectant distribution.
- Published
- 1996
22. Measles virus induction of human endothelial cell tissue factor procoagulant activity in vitro.
- Author
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Mazure G, Grundy JE, Nygard G, Hudson M, Khan K, Srai K, Dhillon AP, Pounder RE, and Wakefield AJ
- Subjects
- Animals, Antibodies, Capsid analysis, Capsid biosynthesis, Cells, Cultured, Chlorocebus aethiops, Cytomegalovirus physiology, Endothelium, Vascular drug effects, Endothelium, Vascular virology, Factor VIII metabolism, Herpesvirus 1, Human physiology, Humans, Immunohistochemistry, Interleukin-1 immunology, Interleukin-1 pharmacology, Kinetics, Lipopolysaccharides pharmacology, Measles virus isolation & purification, Measles virus radiation effects, Recombinant Proteins pharmacology, Thromboplastin analysis, Time Factors, Ultraviolet Rays, Umbilical Veins, Vero Cells, Viral Core Proteins analysis, Viral Core Proteins biosynthesis, Endothelium, Vascular physiology, Measles virus physiology, Thromboplastin biosynthesis
- Abstract
Measles virus infection of microvascular endothelium in vivo and ensuing endothelial cell activation may be important in the pathogenesis of subsequent inflammation in target organs. This study investigated the capacity of measles virus to induce procoagulant activity, in vitro, in endothelial cells isolated from human umbilical cord veins. Endothelial cells were infected with a clinical isolate of measles virus propagated in Vero cells. Cells were also incubated with bacterial lipopolysaccharide (10 micrograms/ml), herpes simplex virus type 1, cytomegalovirus or culture medium alone as positive and negative controls, respectively. Endothelial cell procoagulant activity was measured in a one-stage clotting assay. Measles virus stimulated both a time and dose-dependent endothelial cell procoagulant response by the induction of tissue factor synthesis, confirmed by both immunocytochemistry and its dependence on factor VII for activity. This activity was reduced by u.v.-irradiation of the virus. Infected cells were analysed by double immunofluorescent staining for both tissue factor and measles virus N-protein, and examined using confocal scanning laser microscopy. Cells expressing tissue factor were also positive for the measles virus N-protein. Low levels of interleukin-1 were detected in some viral inocula derived from measles virus-infected Vero cells, however neutralising antibody to interleukin-1 failed to inhibit the endothelial cell procoagulant response to measles virus, whereas it significantly reduced procoagulant activity induced in endothelial cells by recombinant interleukin-1. The capacity of measles virus to induce endothelial tissue factor in vitro, may be relevant to the thrombotic vasculopathy associated with measles virus infection in vivo.
- Published
- 1994
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23. Urea and salt effects on enzymes from estivating and non-estivating amphibians.
- Author
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Grundy JE and Storey KB
- Subjects
- Animals, Estivation physiology, Glycolysis physiology, Potassium Chloride pharmacology, Anura metabolism, Cations pharmacology, Methylamines pharmacology, Phosphofructokinase-1 drug effects, Pyruvate Kinase drug effects, Rana pipiens metabolism, Urea pharmacology
- Abstract
The effects of urea, cations (K+,NH4,Na+,Cs+,Li+), and trimethylamines on the maximal activities and kinetic properties of pyruvate kinase (PK) and phosphofructokinase (PFK) from skeletal muscle were analyzed in two anuran amphibians, an estivating species, the spadefoot toad Scaphiopus couchii, and a semi-aquatic species, the leopard frog Rana pipiens. Urea, which accumulates naturally to levels of 200-300 mM during estivation in toads, had only minor effects on the Vmax, kinetic constants and pH curves of PK from either species and no effects on PFK Vmax or kinetic constants. Trimethylamine oxide neither affected enzyme activity directly or changed enzyme response to urea. By contrast, high KCl (200 mM) lowered the Vmax of toad PFK and of PK from both species and altered the Km values for both substrates of frog PFK. Other cations were even more inhibitory; for example, the Vmax of PK from either species was reduced by more than 80% by the addition of 200 mM NH4Cl, NaCl, CsCi, or LiCl. High KCl also significantly changed the Km values for substrates of toad lactate dehydrogenase and strongly reduced the Vmax of glutamate dehydrogenase and NAD-dependent isocitrate dehydrogenase in both species whereas 300 mM urea had relatively little effect on these enzymes. The perturbing effect of urea on enzymes and the counteracting effect of trimethylamines that has been reported for elasmobranch fishes (that maintain high concentrations of both solutes naturally) does not appear to apply to amphibian enzymes.(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1994
- Full Text
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24. Sequence variation within neutralizing epitopes of the envelope glycoprotein B of human cytomegalovirus: comparison of isolates from renal transplant recipients and AIDS patients.
- Author
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Roy DM, Grundy JE, and Emery VC
- Subjects
- AIDS-Related Opportunistic Infections microbiology, Amino Acid Sequence, Base Sequence, Humans, Kidney Transplantation immunology, Molecular Sequence Data, Neutralization Tests, Opportunistic Infections microbiology, Viral Envelope Proteins immunology, Viral Vaccines genetics, Cytomegalovirus genetics, DNA, Viral genetics, Epitopes genetics, Genetic Variation genetics, Viral Envelope Proteins genetics
- Abstract
The envelope glycoprotein B of human cytomegalovirus (CMV) is a major target of the neutralizing antibody response against this virus, and hence has importance as a potential subunit vaccine. PCR was utilized to amplify DNA encoding the dominant antigenic determinant on this molecule, AD-1 (codons 552 to 635), and DNA sequencing was carried out in order to compare nucleotide variation in AD-1 between clinical isolates of CMV and the laboratory strain AD169. Wild-type CMV strains isolated from AIDS patients were not only more likely to possess nucleotide substitutions (19/24 compared to 5/25, P < 0.0001) than those from renal transplant recipients, but they also exhibited a greater degree of nucleotide sequence divergence (6.94 versus 0.82 substitutions/1000 bp, P < 0.0001; 96.0 to 100% versus 99.4 to 100% similarity). Increased sequence variation in the AIDS patients did not correlate with absolute peripheral blood CD4+ T cell level (r = 0.33, P > 0.1). Only two strains from AIDS patients and one strain from the renal transplant recipients possessed nucleic acid substitutions that resulted in codon changes, indicating that AD-1 is relatively well conserved amongst clinical isolates of CMV. The demonstration of strains with codon changes within neutralizing epitopes, however, highlights the importance of taking into consideration the presence of these strains within the wild-type virus population when preparing subunit vaccines.
- Published
- 1993
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25. Transfer of humoral immunity against cytomegalovirus proteins following transplantation of T-cell-depleted allogeneic bone marrow from seropositive donors.
- Author
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Roy DM, Brenner MK, Cook D, Duncan RI, Griffiths PD, and Grundy JE
- Subjects
- Bone Marrow Cells, Cell Line, Chi-Square Distribution, Humans, Immunoblotting, Viral Proteins immunology, Antibodies, Viral biosynthesis, Bone Marrow Transplantation immunology, Cytomegalovirus immunology, Cytomegalovirus Infections immunology, T-Lymphocytes immunology
- Abstract
Previous work by Grob et al. [Lancet i: 774, 1987] has demonstrated that allogeneic, T-cell-depleted bone marrow transplant recipients have a better prognosis for reactivated cytomegalovirus (CMV) infection if their donor is also immune. It was proposed that adoptively transferred humoral immunity was responsible for the protective effect of active infection. Immunoblot analysis using purified virions was used here to examine pre- and posttransplant antibody responses of seropositive recipients who had undergone active viral infection after transplantation. Immunoblots were assessed for the numbers of polypeptides recognised and reactivity against individual polypeptides. Immunoblots were also scanned by quantitative densitometry, and the intensity of antibody responses against total viral protein and individual polypeptides was determined. Sera from recipients with immune donors exhibited a secondary-type immune response in terms of both intensity and polypeptide specific pattern of antibody reactivity, compared with those recipients with nonimmune donors. In particular, recipients with immune donors appeared to show a greater reactivity against a protein of M(r) 55,000; this may represent the envelope glycoprotein gB, which is a major target for neutralising antibodies, and might also be utilised for preparing an effective vaccine for CMV.
- Published
- 1993
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26. The significance of low bcl-2 expression by CD45RO T cells in normal individuals and patients with acute viral infections. The role of apoptosis in T cell memory.
- Author
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Akbar AN, Borthwick N, Salmon M, Gombert W, Bofill M, Shamsadeen N, Pilling D, Pett S, Grundy JE, and Janossy G
- Subjects
- Acquired Immunodeficiency Syndrome immunology, Acute Disease, Adult, Apoptosis drug effects, Apoptosis genetics, Biopsy, Cells, Cultured, Chickenpox immunology, Female, Fibroblasts physiology, Humans, Infectious Mononucleosis immunology, Interleukin-2 physiology, Lymph Nodes immunology, Lymph Nodes pathology, Lymphocyte Activation, Male, Middle Aged, Protein-Tyrosine Kinases genetics, Proto-Oncogene Proteins genetics, Proto-Oncogene Proteins c-bcl-2, T-Lymphocyte Subsets metabolism, Apoptosis physiology, Immunologic Memory physiology, Leukocyte Common Antigens, Proto-Oncogene Proteins biosynthesis, T-Lymphocyte Subsets immunology, Virus Diseases immunology
- Abstract
The bcl-2 gene product has been shown to prevent apoptotic cell death. We have now investigated the bcl-2 protein expression by resting and activated mature T cell populations. Freshly isolated CD45RO+ T cells within CD4+ and CD8+ subsets expressed significantly less bcl-2 than CD45RO- (CD45RA+) T cells (p < 0.001). When CD45RA+ T cells within both CD4+ and CD8+ subsets were activated in vitro, the transition to CD45RO phenotype was associated with a decrease in bcl-2 expression. Patients with acute viral infections such as infectious mononucleosis caused by Epstein-Barr virus infections or chickenpox, resulting from varicella zoster virus infection, had circulating populations of activated CD45RO+ T cells which also showed low bcl-2 expression. In these patients, a significant correlation was seen between low bcl-2 expression by activated T cells and their apoptosis in culture (r = 0.94, p < 0.001). These results suggest that the primary activation of T cells leads to the expansion of a population that is destined to perish unless rescued by some extrinsic event. Thus the suicide of CD45RO+ T cells could be prevented by the addition of interleukin 2 to the culture medium which resulted in a concomitant increase in the bcl-2 expression of these cells. Alternatively, apoptosis was also prevented by coculturing the activated T lymphocytes with fibroblasts, which maintained the viability of lymphoid cells in a restinglike state but with low bcl-2 expression. The paradox that the CD45RO+ population contains the primed/memory T cell pool yet expresses low bcl-2 and is susceptible to apoptosis can be reconciled by the observations that maintenance of T cell memory may be dependent on the continuous restimulation of T cells, which increases their bcl-2 expression. Furthermore, the propensity of CD45RO+ T cells to extravasate may facilitate encounter with fibroblast-like cells in tissue stroma and thus be an important additional factor which promotes the survival of selected primed/memory T cells in vivo.
- Published
- 1993
- Full Text
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27. Increased adherence of CD2 peripheral blood lymphocytes to cytomegalovirus-infected fibroblasts is blocked by anti-LFA-3 antibody.
- Author
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Grundy JE, Pahal GS, and Akbar AN
- Subjects
- Antibodies, Monoclonal immunology, Binding, Competitive, CD2 Antigens, CD58 Antigens, Cell Adhesion immunology, Cell Adhesion Molecules immunology, Cells, Cultured, Fibroblasts immunology, Humans, Intercellular Adhesion Molecule-1, Leukocytes immunology, Antigens, CD immunology, Antigens, Differentiation, T-Lymphocyte analysis, Cytomegalovirus Infections immunology, Membrane Glycoproteins immunology, Receptors, Immunologic analysis, T-Lymphocytes immunology
- Abstract
In the accompanying manuscript (p. 405) we describe the up-regulation of the adhesion molecules LFA-3 and ICAM-1 on the surface of cytomegalovirus (CMV)-infected fibroblasts from days 1 to 5 post-infection. Peak expression was seen on day 2, when LFA-3 was twice, and ICAM-1 three times, the level on uninfected fibroblasts. The present study demonstrates a parallel increase in the adhesion of peripheral blood leucocytes to the CMV-infected fibroblasts, which was significantly greater than adhesion to uninfected fibroblasts from days 2 to 4 post-infection. This effect was seen from 2 to 24 hr of leucocyte-fibroblast co-culture. The increased binding to infected fibroblasts was accounted for by the CD2+ subset of lymphocytes. All subpopulations of CD2+ lymphocytes, namely CD3+, CD4+, and CD8+ cells, demonstrated increased adhesion to CMV-infected fibroblasts, suggesting that the CD2-LFA-3 interaction was an important component of the increased binding. This proposal was supported by the fact that the pretreatment of infected fibroblasts with monoclonal antibodies specific for LFA-3, significantly blocked the binding of CD2+ lymphocytes. Supernatants from infected fibroblasts, or co-cultures of leucocytes and infected fibroblasts, could transfer increased leucocyte binding to uninfected fibroblasts, suggesting that CMV might accentuate inflammatory responses. As lymphocytes can be activated by the CD2 pathway, CMV might also provoke nonspecific leucocyte responses to uninfected as well as infected cells, which could possibly contribute to tissue damage.
- Published
- 1993
28. Up-regulation of LFA-3 and ICAM-1 on the surface of fibroblasts infected with cytomegalovirus.
- Author
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Grundy JE and Downes KL
- Subjects
- Antigens, Surface analysis, CD58 Antigens, Cells, Cultured, Fibroblasts immunology, Fluorescent Antibody Technique, Histocompatibility Antigens Class I analysis, Humans, Intercellular Adhesion Molecule-1, Antigens, CD analysis, Cell Adhesion Molecules analysis, Cytomegalovirus Infections immunology, Membrane Glycoproteins analysis
- Abstract
The effect of cytomegalovirus (CMV) infection on the cell surface expression of the adhesion molecules LFA-3 and ICAM-1 was studied by flow cytometry using human embryo lung fibroblasts. Our results demonstrated a marked increase in the expression of these molecules from days 1 to 5 post-infection. Peak expression of LFA-3 and ICAM-1 on the surface of the infected cell occurred at 2 days post-infection, when LFA-3 was twofold, and ICAM-1 threefold, greater than the level observed on uninfected fibroblasts. In contrast, parallel studies on the expression of class I HLA confirmed our previous findings that CMV induces a down-regulation of this molecule on the surface of the infected cell, and further demonstrated that at the time of maximum increase in LFA-3 and ICAM-1 expression, class I HLA expression was only 46% of the uninfected cell level. Immunofluorescence and confocal scanning laser microscopy revealed markedly enhanced expression of LFA-3 in infected cells, with accumulations in discrete granules in the perinuclear area, contrasting with the diffuse cytoplasmic distribution of this molecule in uninfected fibroblasts. ICAM-1 was found to be highly localized at the cell membrane of infected cells, whereas a diffuse cytoplasmic distribution was observed in the uninfected cell. The mechanism of the up-regulation of adhesion molecule expression on the CMV-infected cells remains to be determined; however cytokines known to up-regulate ICAM-1 were not detected in the culture supernatant of infected cells. The effects of CMV infection on the adhesion of peripheral blood leucocytes to fibroblasts is described in the accompanying manuscript (p. 413).
- Published
- 1993
29. Evaluation of neutralizing antibody titers against human cytomegalovirus in intravenous gamma globulin preparations.
- Author
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Roy DM and Grundy JE
- Subjects
- Humans, Neutralization Tests, Antibodies, Viral analysis, Cytomegalovirus immunology, Immunoglobulins, Intravenous chemistry
- Published
- 1992
30. Down-regulation of the class I HLA heterodimer and beta 2-microglobulin on the surface of cells infected with cytomegalovirus.
- Author
-
Barnes PD and Grundy JE
- Subjects
- Antigens, Surface isolation & purification, Biological Transport, Cell Compartmentation, Cell Membrane Permeability, Cells, Cultured, Down-Regulation, Fibroblasts, Fluorescent Antibody Technique, HLA Antigens isolation & purification, Histocompatibility Antigens Class I isolation & purification, Humans, Macromolecular Substances, beta 2-Microglobulin isolation & purification, Antigens, Surface metabolism, Cytomegalovirus metabolism, HLA Antigens metabolism, Histocompatibility Antigens Class I metabolism, beta 2-Microglobulin metabolism
- Abstract
Cytotoxic T cell recognition of virus-infected cells requires the presentation of viral peptides by class I HLA molecules on the cell surface. We report here that cytomegalovirus (CMV) infection of human fibroblasts results in a progressive decrease in the cell surface expression of class I HLA and beta 2-microglobulin (beta 2m) such that in the late stages of infection the majority of infected cells have no detectable surface class I HLA. Coincident with decreased surface expression of class I HLA was an increase in his cytoplasmic expression. Confocal scanning laser microscopic analysis demonstrated that class I HLA and beta 2m accumulate in a perinuclear compartment inside the CMV-infected cell. Our data thus support the concept that CMV infection induces altered transport of class I HLA to the cell surface. We suggest that the virus has evolved this mechanism as a strategy to avoid T cell recognition of infected cells.
- Published
- 1992
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31. Severe cytomegalovirus pneumonitis in HIV infected patients with higher than average CD4 counts.
- Author
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Squire SB, Lipman MC, Bagdades EK, Mulvenna PM, Grundy JE, Griffiths PD, and Johnson MA
- Subjects
- Adult, Cytomegalovirus Infections immunology, HIV Infections immunology, Humans, Male, Opportunistic Infections immunology, Pneumonia, Viral immunology, CD4 Antigens analysis, Cytomegalovirus Infections complications, HIV Infections complications, Opportunistic Infections complications, Pneumonia, Viral complications
- Abstract
Background: Cytomegalovirus may replicate within the lungs both of recipients of transplants and of patients infected with the human immunodeficiency virus (HIV). A hypothesis formulated by this group was that a host damaging immune response might be provoked by cytomegalovirus infection and cause a severe pneumonitis in recipients of allogeneic transplants, whereas the progressive impairment of cellular immunity in patients with HIV disease would preclude a damaging immune response in the lungs, and thus protect these patients from severe cytomegalovirus pneumonitis. This study set out to discover whether severe cytomegalovirus pneumonitis arises in HIV infected patients., Methods: Data were prospectively collected on severity of pneumonitis and infectious agents identified in consecutive respiratory episodes in HIV infected patients undergoing diagnostic bronchoalveolar lavage during 20 months., Results: Eighty five episodes of pneumonitis occurred in 68 patients. Cytomegalovirus was identified as the only infectious agent in nine episodes (nine patients). Seven of the episodes were mild; all these patients had CD4 counts below 0.1 x 10(9)/1. The remaining two episodes were severe and ventilatory support was required. In both cases the CD4 counts were above 0.2 x 10(9)/1 and HIV infection appeared to have been acquired shortly before presentation., Conclusion: Although rare, severe cytomegalovirus pneumonitis may occur in HIV infected patients. Both patients with severe pneumonitis in this series had relatively well preserved immune function. These findings support the hypothesis that severe cytomegalovirus pneumonitis is an immunopathological condition.
- Published
- 1992
- Full Text
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32. Beta 2 microglobulin on the envelope of urinary cytomegalovirus is not associated with host class I human leukocyte antigen alpha chain.
- Author
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Rose JS and Grundy JE
- Subjects
- Antibodies, Monoclonal immunology, Cytomegalovirus Infections microbiology, Epitopes immunology, HLA Antigens immunology, Humans, beta 2-Microglobulin immunology, Cytomegalovirus chemistry, Genes, MHC Class I, HLA Antigens analysis, Urine microbiology, beta 2-Microglobulin analysis
- Abstract
Previous studies have shown that beta 2 microglobulin (beta 2m) is associated with glycoproteins present on the envelope of urinary human cytomegalovirus (CMV). beta 2m is non-covalently associated with the alpha chain of human leukocyte antigen (HLA) class I antigens and therefore it was of interest to determine whether the class I alpha chain is also associated with the beta 2m-CMV complex. Using a panel of monoclonal antibodies recognizing different conformational determinants on the HLA class I heterodimer or free beta 2m, we have shown that beta 2m but not the alpha chain of HLA class I could be immunoprecipitated from 125I-surface-labelled virions purified directly from urine. We therefore conclude that host class I HLA alpha chains are not associated with beta 2m on the envelope of urinary CMV.
- Published
- 1992
- Full Text
- View/download PDF
33. Double-blind, placebo-controlled trial of human lymphoblastoid interferon prophylaxis of cytomegalovirus infection in renal transplant recipients.
- Author
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Lui SF, Ali AA, Grundy JE, Fernando ON, Griffiths PD, and Sweny P
- Subjects
- Adult, Cytomegalovirus isolation & purification, Cytomegalovirus Infections etiology, Cytomegalovirus Infections microbiology, Double-Blind Method, Female, Humans, Interferon-alpha administration & dosage, Interferon-alpha adverse effects, Male, Pneumonia, Viral prevention & control, Cytomegalovirus Infections prevention & control, Interferon-alpha pharmacology, Kidney Transplantation adverse effects
- Abstract
We have conducted a double-blind, placebo-controlled trial of human lymphoblastoid interferon prophylaxis of cytomegalovirus (CMV) infection in 74 renal transplant recipients. Interferon (3 x 10(6) units was given thrice weekly for the first 6 weeks, then twice weekly for a further 8 weeks. During the period of interferon therapy, the incidence of CMV excretion was lower in the interferon group (28% versus 50%, P = 0.065), mainly due to a significant reduction of CMV reactivation (9% versus 56%, P = 0.02). However, for the whole study period (including the follow-up period after interferon therapy), there was no difference in the incidence of CMV excretion (44% versus 53%). The onset of CMV excretion was delayed (8.2 +/- 0.8 to 20.9 +/- 5.5 weeks, P = 0.04). The duration of CMV excretion was also reduced (11.1 +/- 3.1 to 29.4 +/- 5.7 weeks, P = 0.008). The number of positive CMV isolates from urine and saliva was significantly less in the interferon group. There was no difference in the site of CMV excretion. Of the patients in the treatment group who excreted CMV, 43% developed disease as compared to 63% in the control group (difference not significant). There was also no significant difference in the severity of the CMV infection between the two groups. Benefit appears to be restricted to seropositive recipients of seronegative kidneys. The interferon regime used in this study was well tolerated, with mild fever being the only reported side-effect. No patient had to stop therapy because of toxicity. The incidence of rejection and graft loss was not different between the two groups.
- Published
- 1992
- Full Text
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34. Use of the polymerase chain reaction to analyse sequence variation within a major neutralizing epitope of glycoprotein B (gp58) in clinical isolates of human cytomegalovirus.
- Author
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Darlington J, Super M, Patel K, Grundy JE, Griffiths PD, and Emery VC
- Subjects
- Amino Acid Sequence, Base Sequence, Blotting, Southern, Bone Transplantation, Cells, Cultured, Cytomegalovirus immunology, DNA, Viral, Humans, Kidney Transplantation, Molecular Sequence Data, Neutralization Tests, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Sequence Homology, Nucleic Acid, Viral Envelope Proteins immunology, Antigenic Variation genetics, Cytomegalovirus genetics, Epitopes genetics, Viral Envelope Proteins genetics
- Abstract
The heterogeneity of low passage human cytomegalovirus (HCMV) strains was determined by HindIII typing of 28 clinical isolates from transplant patients. These data have shown that, in general, each patient's strain has a unique restriction profile, usually comprising combinations of HindIII sites present in one or more of the tissue culture-adapted strains AD169, Towne and Davis. To map sequence changes in a more refined manner we performed detailed analyses of 33 low passage clinical isolates, including those aforementioned, analysing a sequence within glycoprotein B containing a major neutralizing epitope. A 149 bp sequence containing the epitope (amino acids 608 to 625) was amplified using the polymerase chain reaction, the products were cloned and their DNA sequence was determined. Comparison of the DNA and deduced amino acid sequences with those of HCMV strain AD169 revealed that there was a high degree of conservation of the epitope between the 33 clinical isolates. However 10 of the isolates possessed silent mutations and three isolates contained mutations producing amino acid changes within the neutralizing epitope. The possible functional significance of these changes is discussed.
- Published
- 1991
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35. Molecular comparisons of the beta 2-microglobulin-binding site in class I major-histocompatibility-complex alpha-chains and proteins of related sequences.
- Author
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Tysoe-Calnon VA, Grundy JE, and Perkins SJ
- Subjects
- Amino Acid Sequence, Animals, Binding Sites, Computer Graphics, Humans, Macromolecular Substances, Models, Molecular, Molecular Sequence Data, Protein Conformation, Sequence Homology, Nucleic Acid, beta 2-Microglobulin immunology, Antigens, CD physiology, Histocompatibility Antigens Class I physiology, beta 2-Microglobulin metabolism
- Abstract
beta 2-Microglobulin (beta 2m) binds non-covalently to the alpha 1, alpha 2 and alpha 3 domains of the alpha-chain of Class I major-histocompatibility-complex (MHC) molecules. On the basis of the crystal structures of human leucocyte antigens HLA-A2.1 and HLA-Aw68.1, we have used molecular-graphics analyses to define 44 contact points between 19 alpha-chain residues and 18 beta 2m residues. In 88 other alpha-chain sequences from the HLA-A, HLA-B, HLA-C, HLA-D, HLA-E, HLA-F and HLA-G locus products in man and the H-2, Qa and Tla loci in mouse, 37 contact sites were conserved to 90% or more, and in beta 2m sequences from seven other species 40% of contact sites were totally conserved. Four distinct regions form the contact points between the alpha-chain and beta 2m, one on each of the alpha 1 and alpha 2 domains and two on the alpha 3 domain. We have further studied the alpha-chain sequences of three non-MHC molecules, human CD1 and rat Fc receptor (FcRn), known to bind to beta 2m, and a third molecule, the putative product of the H301 (UL18) gene of human cytomegalovirus (CMV). CMV has been shown to bind beta 2m, and it has been postulated that the H301-gene product, which has sequence similarity to Class I HLA, is the protein responsible. These sequences exhibited much lower residue conservation with the MHC-linked group, although the alpha 3 domain was the most highly conserved, and gaps and insertions were required for optimal alignments with the 90 alpha-chain sequences. Of the 44 beta 2m-alpha-chain contacts defined for Class I HLA, 24 alpha-chain contact sites were conserved in CD1, 25 in FcRn and 17 in the H301-gene product. For CD1 and FcRn, the majority of the conserved beta 2m contacts were found in the alpha 2 domain and the major contact region in the alpha 3 domain. Together with the use of secondary-structure predictions, it was concluded that the binding of beta 2m in CD1 and FcRn was MHC-like at the alpha 3 domain, and probably also at the alpha 2 domain for FcRn, but non-MHC-like for the alpha 1 domain of both molecules and the alpha 2 domain of CD1. In the H301-gene product sequence, only the beta 2m contacts with the main region of the alpha 3 domain were noticeably conserved.(ABSTRACT TRUNCATED AT 400 WORDS)
- Published
- 1991
- Full Text
- View/download PDF
36. Alterations of cellular proteins in human cytomegalovirus infection: potential for disease pathogenesis.
- Author
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Grundy JE
- Subjects
- Humans, Protein Binding, Cytomegalovirus physiology, Cytomegalovirus Infections physiopathology, HLA Antigens physiology, beta 2-Microglobulin metabolism
- Published
- 1991
37. IL2 activated killer cells may contribute to cytomegalovirus induced marrow hypoplasia after bone marrow transplantation.
- Author
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Duncombe AS, Grundy JE, Prentice HG, and Brenner MK
- Subjects
- Adolescent, Adult, Antibodies, Monoclonal immunology, Antibodies, Monoclonal physiology, Antibodies, Monoclonal therapeutic use, Cells, Cultured, Child, Cytomegalovirus Infections drug therapy, Cytomegalovirus Infections pathology, Cytotoxicity, Immunologic, Humans, Interleukin-2 biosynthesis, Macrophages immunology, Major Histocompatibility Complex, Monocytes metabolism, Pneumonia, Viral pathology, Receptors, Interleukin-2 immunology, Bone Marrow pathology, Bone Marrow Transplantation, Cytomegalovirus physiology, Cytomegalovirus Infections etiology, Killer Cells, Lymphokine-Activated physiology, Pneumonia, Viral etiology
- Abstract
Cytomegalovirus (CMV) infection remains the commonest cause of infective death following allogeneic bone marrow transplantation (BMT). CMV disease post-BMT occurs in the context of compromised cellular defence mechanisms and is associated with marrow hypoplasia and pneumonitis. However, CMV infection induces release of interleukin 2 (IL2) which in turn generates MHC unrestricted lymphokine activated killer (LAK) cells. We have investigated whether recruitment of IL2 activated MHC unrestricted defence mechanisms post-transplant can be implicated in the marrow hypoplasia that frequently accompanies CMV infection. The results show that IL2 activated peripheral blood mononuclear cells (PBMC) have substantial cytotoxicity against MHC matched and MHC mismatched marrow fibroblasts but that this activity is not specific for CMV infected fibroblasts; uninfected target cells are also equally killed. Furthermore, post-BMT PBMC show greater responsiveness to IL2 than normal PBMC in killing of marrow fibroblasts. We provide a hypothesis from these observations which may explain some of the consequences of CMV infection post-BMT. Local production of IL2 activated cytotoxic cells which would be generated during CMV infection would damage uninfected as well as infected marrow fibroblasts and thereby could compromise haemopoietic growth factor production by marrow fibroblasts. Similarly generated cytotoxicity in the lung may accompany CMV pneumonitis. Our results suggest that administration of anti-IL2 receptor antibody may have a therapeutic role in CMV disease post-BMT as has recently been shown in graft-versus-host disease.
- Published
- 1991
38. CMV-induced augmentation of GVL effect may be mediated by cytokines.
- Author
-
Duncombe AS, Grundy JE, Prentice HG, and Brenner MK
- Subjects
- Humans, Interferon-gamma immunology, Lymphocytes metabolism, Tumor Necrosis Factor-alpha immunology, Tumor Necrosis Factor-alpha metabolism, Bone Transplantation immunology, Cytokines immunology, Cytomegalovirus immunology, Graft Rejection immunology
- Published
- 1991
39. High conservation of beta 2-microglobulin contact residues among 82 class I major histocompatability complex alpha-chain sequences.
- Author
-
Tysoe-Calnon VA, Grundy JE, Nealis AS, and Perkins SJ
- Subjects
- Animals, Biological Evolution, Genes, MHC Class I, Genetic Variation, H-2 Antigens genetics, HLA Antigens genetics, Macromolecular Substances, Major Histocompatibility Complex, Mice, Histocompatibility Antigens Class I genetics, beta 2-Microglobulin genetics
- Published
- 1990
- Full Text
- View/download PDF
40. Virologic and pathogenetic aspects of cytomegalovirus infection.
- Author
-
Grundy JE
- Subjects
- Cytomegalovirus immunology, Cytomegalovirus Infections immunology, Cytomegalovirus Infections microbiology, Graft Rejection, Humans, Lung microbiology, Pneumonia, Viral immunology, Pneumonia, Viral microbiology, Cytomegalovirus pathogenicity, Cytomegalovirus Infections etiology, Pneumonia, Viral etiology
- Abstract
Cytomegalovirus (CMV) is a ubiquitous agent that rarely causes disease in immunocompetent humans but is an important cause of morbidity and mortality in the immunocompromised. A number of host and viral factors are associated with pathology following CMV infection. CMV can be found at many sites in the body but only causes disease at some of these and then only in certain patient populations. In some situations the mechanism underlying disease is direct viral replication, but in others, particularly CMV pneumonitis in allogeneic transplant recipients, an immunopathologic basis is strongly implicated. An important factor in the pathogenesis of infection and the expression of symptomatic disease is the source of CMV infection-whether it arises from an exogenous source or is due to reactivation of latent endogenous virus. Exogenous infection can occur in previously seronegative individuals or in those with prior exposure to the virus. In renal transplant recipients both types of exogenous infection have been associated with disease. Another factor that affects the interaction between CMV and its host is the modulating effect of the virus on the host immune response, the mechanism of which remains unknown.
- Published
- 1990
- Full Text
- View/download PDF
41. Cytomegalovirus in the lungs of patients with AIDS. Respiratory pathogen or passenger?
- Author
-
Millar AB, Patou G, Miller RF, Grundy JE, Katz DR, Weller IV, and Semple SJ
- Subjects
- Antibodies, Monoclonal, Bronchoalveolar Lavage Fluid microbiology, Cytomegalovirus isolation & purification, Cytomegalovirus Infections diagnosis, Humans, Pneumonia, Viral diagnosis, Acquired Immunodeficiency Syndrome complications, Cytomegalovirus pathogenicity, Cytomegalovirus Infections complications, Lung microbiology, Pneumonia, Viral complications
- Abstract
A total of 166 consecutive clinical episodes of pneumonitis in patients with acquired immune deficiency syndrome (AIDS) or antibody positive for human immune deficiency virus (HIV) were investigated for evidence of cytomegalovirus (CMV) infection in their lungs and at peripheral sites to determine the pathogenicity of this virus in the lung and its relationship to peripheral CMV shedding. Evidence of CMV infection was sought in bronchoalveolar lavage (BAL) fluid, blood, saliva, and urine using a specific monoclonal antibody to antigens produced by CMV-infected cells within 24 h. Although CMV was detected in 31 (19%) of BAL fluid specimens, in only six episodes was this the sole pathologic finding. In the remaining episodes either another infectious agent, Kaposi's sarcoma, or lymphoid interstitial pneumonitis was found or no pathogen was detected. None of the patients were given specific anti-CMV treatment, and all but two recovered, including those patients in whom CMV was the sole finding at BAL. The presence of peripheral shedding of CMV did not have any significance in mortality or morbidity. Our findings are in direct contrast to those in recipients of allogeneic bone marrow transplants, in whom CMV pneumonitis is associated with a high mortality. We postulate that this difference is because AIDS patients cannot mount the destructive immune response to CMV in the lung, which we believe to be the basis of the pathology seen in the former group. We conclude that CMV is not a pathogen in the lungs of patients with HIV infection, and we suggest that its presence at this site does not warrant specific therapy in these patients.(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1990
- Full Text
- View/download PDF
42. Role of beta 2-microglobulin in cytomegalovirus infection.
- Author
-
Grundy JE
- Subjects
- Carrier Proteins metabolism, Cytomegalovirus metabolism, Cytomegalovirus Infections genetics, Cytomegalovirus Infections immunology, Genetic Predisposition to Disease, Histocompatibility Antigens Class I metabolism, Humans, Receptors, Virus metabolism, Viral Proteins metabolism, beta 2-Microglobulin metabolism, Cytomegalovirus Infections metabolism, beta 2-Microglobulin physiology
- Published
- 1990
- Full Text
- View/download PDF
43. Detection of cytomegalovirus by ELISA in urine samples is inhibited by beta 2 microglobulin.
- Author
-
McKeating JA, Grundy JE, Varghese Z, and Griffiths PD
- Subjects
- Antigens, Viral analysis, Cells, Cultured, Chromatography, Gel, Cytomegalovirus immunology, Enzyme-Linked Immunosorbent Assay, Epitopes immunology, Humans, Muramidase pharmacology, beta 2-Microglobulin isolation & purification, Cytomegalovirus isolation & purification, Urine microbiology, beta 2-Microglobulin urine
- Abstract
During development of an enzyme immunoassay for the detection of cytomegalovirus (CMV) we previously discovered that virus found naturally in urine specimens could not be captured onto the solid phase by CMV-specific monoclonal antibodies, whereas these same antibodies could capture CMV from cell culture supernatants. We now report that urine from normal CMV-seronegative individuals contains a substance of molecular weight 11-12,000 daltons that inhibits the ELISA detection of cell culture-grown CMV. The addition of a known urinary protein of this molecular weight, beta 2 microglobulin (beta 2m; 11,700 daltons), inhibited the detection of cell culture-grown CMV in the ELISA over the concentration range found in clinical urine samples. In contrast, another low molecular weight urinary protein, lysozyme, had no inhibitory effect. beta 2m caused inhibition only when added to the virus preparation and not to the antibody-capture stage. We conclude that beta 2m in urine prevents the detection of CMV by ELISA by binding to the virus and masking its antigenic determinants and we calculate that of the order of 10(5) molecules of beta 2m bind to each particle of cell culture-grown CMV. We postulate that CMV in fresh urine specimens is similarly coated with beta 2m, accounting for the failure to detect it by ELISA.
- Published
- 1986
- Full Text
- View/download PDF
44. Effect of murine cytomegalovirus infection on mitogen responses in genetically resistant and susceptible mice.
- Author
-
Allan JE, Shellam GR, and Grundy JE
- Subjects
- Animals, B-Lymphocytes, Cell Count, Cytomegalovirus growth & development, Cytomegalovirus Infections genetics, Female, Immunity, Innate, Liver microbiology, Macrophages, Mice, Mice, Inbred BALB C, Mice, Inbred C3H, Organ Size, Spleen microbiology, T-Lymphocytes, Cytomegalovirus Infections immunology, Lymphocyte Activation
- Abstract
Suppression of the blastogenic response of spleen cells was found during murine cytomegalovirus infection of the genetically susceptible BALB/c and also the more resistant BALB.K strains of mice. These results were observed for both the T-cell mitogen concanavalin A and the B-cell mitogen lipopolysaccharide. As the viral inoculum was increased, there was greater immunosuppression within each strain, the time of maximum depression coinciding with peak virus titers in the spleen. Although both strains developed similar splenic virus titers and exhibited a similar decrease in the proportion of splenic T-lymphocytes, there was greater suppression of the mitogenic response during sublethal infection of the more susceptible BALB/c strain. The suppression could not be readily accounted for by the presence of suppressor cells or by a change in sensitivity to mitogen. The results suggest that the extent of immunosuppression induced by murine cytomegalovirus is determined in part by host genotype.
- Published
- 1982
- Full Text
- View/download PDF
45. Cytomegalovirus infection and progression to AIDS.
- Author
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Webster A, Grundy JE, Lee CA, Emery VC, Cook DG, Kernoff PB, and Griffiths PD
- Subjects
- Humans, Acquired Immunodeficiency Syndrome prevention & control, Acyclovir therapeutic use, Cytomegalovirus Infections drug therapy
- Published
- 1989
- Full Text
- View/download PDF
46. The effect of cytomegalovirus infection on the host response to foreign and hapten-modified self histocompatibility antigens.
- Author
-
Grundy JE and Shearer GM
- Subjects
- Animals, Antibody Formation, Autoantibodies immunology, Immune Tolerance, Mice, Mice, Inbred Strains, Spleen cytology, Time Factors, Cytomegalovirus Infections immunology, Haptens immunology, Histocompatibility Antigens immunology
- Abstract
The effect of murine cytomegalovirus (MCMV) infection on the host's ability to respond to allogeneic or hapten-modified syngeneic histocompatibility antigens was characterized by an early suppressive phase followed by a phase of enhancement. Suppression of the potential to respond to allogeneic H-2 antigens, as measured by an in vitro cell-mediated lympholysis assay, was seen on days 2-4 postinfection with MCMV, and was greatest at larger doses of virus and in mice with H-2 haplotypes associated with genetic susceptibility to MCMV. In mice of the C57BL genetic background, such responses had returned to normal levels by day 6 and thereafter (days 7-13) the potential of the infected host to respond to allogeneic histocompatibility determinants was markedly enhanced compared with uninfected control mice. This enhancement of alloreactivity was maximal at lower doses of virus and was coincident with the emergence of reactivity to self antigens, as measured by autoantibody production. A similar pattern of time-dependent suppression and enhancement of the hapten modified self cytotoxic response was observed. Enhancement of alloreactivity during MCMV infection was dependent on the genetic constitution of the host with the response of BALB/c mice remaining in a suppressive phase for up to 17 days postinfection. The possibility that this in vitro finding of enhanced alloreactivity during MCMV infection underlies the clinical observations of association between episodes of kidney allograft rejection and cytomegalovirus infection is discussed.
- Published
- 1984
- Full Text
- View/download PDF
47. The status of CMV as a human pathogen.
- Author
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Griffiths PD and Grundy JE
- Subjects
- Adult, Animals, Cytomegalovirus genetics, Cytomegalovirus Infections immunology, Disease Models, Animal, Humans, Immune Tolerance, Infant, Infant, Newborn, Oncogenes, Syndrome, Cytomegalovirus pathogenicity, Cytomegalovirus Infections microbiology
- Published
- 1988
- Full Text
- View/download PDF
48. Cytomegalovirus in urine specimens has host beta 2 microglobulin bound to the viral envelope: a mechanism of evading the host immune response?
- Author
-
McKeating JA, Griffiths PD, and Grundy JE
- Subjects
- Antibodies, Monoclonal, Cytomegalovirus immunology, Cytomegalovirus isolation & purification, Cytomegalovirus Infections urine, Humans, Neutralization Tests, Protein Binding, Viral Envelope Proteins isolation & purification, beta 2-Microglobulin immunology, beta 2-Microglobulin urine, Cytomegalovirus metabolism, Cytomegalovirus Infections immunology, Viral Envelope Proteins metabolism, beta 2-Microglobulin metabolism
- Abstract
We have previously reported that human cytomegalovirus (CMV) from urine specimens cannot be captured onto a solid phase by CMV-specific monoclonal antibodies that can capture CMV grown in vitro. We report here that CMV exists in vivo in body fluids such as urine as beta 2 microglobulin (beta 2 m)-coated particles. We have demonstrated the presence of beta 2m on CMV purified directly from urine by Western blotting and have shown that the beta 2m was associated with the viral envelope. Urinary CMV could be specifically bound by an affinity column comprising a monoclonal antibody specific for beta 2m bound to Sepharose. The beta 2m-coated urinary CMV could not be neutralized by hyperimmune globulin, human immune sera or murine monoclonal antibodies that could neutralize CMV grown in cell culture. We conclude that the binding of beta 2m by CMV masks the important antigenic sites necessary for neutralization which are recognized by man's immune response. We propose that CMV has evolved this mechanism of coating itself in a host protein as a mechanism of evading the host immune response and facilitating transmission between individuals.
- Published
- 1987
- Full Text
- View/download PDF
49. Effect of Nu/Nu gene on genetically determined resistance to murine cytomegalovirus.
- Author
-
Grundy JE and Melief CJ
- Subjects
- Animals, Cytomegalovirus Infections genetics, Disease Susceptibility, Lethal Dose 50, Mice, Mice, Inbred BALB C, Mice, Inbred CBA, Cytomegalovirus Infections immunology, Major Histocompatibility Complex, Mice, Nude genetics, T-Lymphocytes immunology
- Abstract
Adult athymic Nu/Nu mice showed increased susceptibility to lethal infection with murine cytomegalovirus (MCMV) when compared to their heterozygous T cell-competent Nu/+ littermates. However, the extent of this increase in susceptibility varied dramatically depending on the genetic background of the mice carrying the Nu/Nu gene. Genetically susceptible Balb/c (H-2d) mice showed a greater than 316-fold difference between the LD50 of Nu/Nu and Nu/+ littermates. In marked contrast, the genetically resistant CBA (H-2K) strain was characterized by only a 16-fold difference in resistance between Nu/Nu and Nu/+ mice, and furthermore, the athymic CBA Nu/Nu mice were no susceptible than the T cell-competent Balb/c Nu/+ strain. These results together with previous observations strongly suggest that the (H-2k)-associated resistance of the CBA strain is mediated by non-T cell-dependent early defence mechanisms.
- Published
- 1982
- Full Text
- View/download PDF
50. The source of cytomegalovirus infection in seropositive renal allograft recipients is frequently the donor kidney.
- Author
-
Grundy JE, Super M, Lui S, Sweny P, and Griffiths PD
- Subjects
- Cytomegalovirus genetics, Cytomegalovirus isolation & purification, Cytomegalovirus Infections diagnosis, Cytomegalovirus Infections immunology, Humans, Prospective Studies, Restriction Mapping, Serotyping, Tissue Donors, Cytomegalovirus Infections etiology, Kidney Transplantation adverse effects
- Published
- 1987
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