bluuebunny/crossref_metadata_2025 huggingface dataset:
Release and Functional Characterization of the Leukotriene D4‐Metabolizing Enzyme (Dipeptidase) from Human Polymorphonuclear Leucocytes
Authors: RAULF M. , KÖNIG W. , KÖLLER M. , STÜNING M.
Published: June 2006

Polymorphonuclear leucocytes released LTD4‐dipeptidase activity in a time‐, calcium‐, and cell number‐dependent fashion. The LTD4‐dipeptidase released from polymorphonuclear leucocytes (PMN) by incubation with calcium (0.91 mm) was detectable up to a cell concentration of 1 × 106/ml and increased with higher concentrations. Maximal LTD4‐dipeptidase activity within the extracellular environment was detected after 15 min of incubation (2x107/ml) in the presence of 2–4.5 mm calcium or after 30 min, when stimulation was carried out with 0.91 mm calcium. The activity of the released LTD4‐dipeptidase was modulated by various metal ions and other compounds. The addition of Mn2+ Co2+, and Zn2+ (final concentration 1 mm) enhanced the LTD4‐dipeptidase activity, while Cu2+ led to a complete inhibition. In the absence of exogenoas calcium EDTA inhibited LTD4‐dipeptidase. Calcium up to a concentration of 5 and 10mm decreased the dipeptidase activity. The LTD4‐dipeptidase is not affected by bestatin, leupeptin. or N‐ethyl‐maleinimide (NEM). The Km of LTD4‐dipeptidase for LTD4 was 0.95±0.2μm and vmax was 737.5±112.5 pmol/min × mg protein (n = 3±SEM). The highest LTD4dipeptidase activity was obtained at physiological pH values. LTD4‐dipeptidase activity can also be released from other cell types, but the enzyme activity from human PMN exceeded that of other cells (e.g. human lymphocytes/monocytes and basophils (LMB) and human lung cell suspension).
The Role of Gamma Interferon in B‐Cell Activation
Authors: PISCHEDDA F. , KIRCHNER H. , LEANDERSON T.
Published: June 2006

This study evaluates the role of gamma interferon in in vitro B‐cell activation. Two elones of an alloreactive helper T‐cell line were equally effective in activating B cells polyclonally to proliferate and to mature to immunoglobulin secretion. One of these clones produced high levels of gamma interferon, while the other clone did not. From these data we conclude that gamma interferon plays no limiting role in T‐dependent B‐cell activation.
Kinetic Analysis of Qa‐1‐Specific Cloned Cytotoxic T Lymphocytes: Lytic Parameters and Evaluation of Cellular Inhibition
Authors: LEFEVER A. V. , TRUITT R. L.
Published: June 2006

A kinetic analysis of lysis assay was used to compare cytolysis of target cells expressing different allelic Qa‐I determinants hy a Qa‐lb‐specific cytotoxic T‐lymphocyte clone (CTL) Although this clone specifically recognized the Qa‐lb dcterminant it also recognized the Qa‐lc and Qa‐ld determinants to a lesser extent. The maximum rate of lysis against Qa‐lb target was l0‐fold faster than against Qa‐lc or ‐Id targets, The affinity of the CTL clone for the Qa‐lb and ‐Ic target cells was the same, but significantly less for Qu‐ld targets. The pattern of inhibition observed for target and inhibitor cells in these experiments was competitive. These studies demonstrated a greater similarity between the Qa‐lb and ‐Ic determinants, compared with Oa‐ld. Kinetic analysis of lytic reactions allowed for quantitative evaluation of the similarities and differences between various target cell populations to an extent not possible using conventional cell‐mediated assays.
Human Alveolar Macrophages Synthesize Active Complement Components C6, C7, and C8 in Vitro
Authors: PETTERSEN H. B. , JOHNSON E. , HETLAND G.
Published: June 2006

We investigated whether serum‐free human alveolar macrophage cultures synthesize aetive C6. C7. and C8. There was a significant binding of polyclonal anti‐human C6 antibodies to agarose beads incubated with unstimulated macrophages for 24 or 48 h. Endotoxin stimulation of the macrophages was necessary for significant binding of polyclonal anti‐C7 and anti‐C8 antibodies to agarose beads co‐cultured for 48 or 96 h. Two monoclonal antibodies (poly C9‐MA and MCaE11) specific for a neoantigen of polymerized C9 in the terminal complement complex (TCC), bound to beads mainly incubated with endotoxin stimulated macrophages. The MCaEll was more sensitive than the poly C9‐MA in detecting the C9 neoantigen on beads incubated with the macrophages or human serum diluted 1:16. We thus conclude that human alveolar macrophages synthesize active C6, C7, and C9 that together with C5 and C9, assemble as the TCC on co‐cultured agarose beads. Activation of the alternative pathway on the agarose with generation of fixed C3 and C5 convertases is a prerequisite for the subsequent generation of the TCC.
Expression of CRl and CR2 Complement Receptors following Epstein‐Barr Virus Infection of Burkitt's Lymphoma Cell Lines
Authors: COHEN J. H. M. , FISCHER E. , KAZATCHKINE M. D. , LENOIR G. M. , LEFEVRE‐DHLVINCOURT C. , REVILLARD J.‐P.
Published: June 2006

Epstein‐Barr virus (EBV) infection of human B lymphocvtes involves a specific receptor closely associated with, or identical to, the C3d complement receptor, CR2, Thus. 25 out of 29 EBV‐positive Burkilt's lymphuma (BL) cell lines but none of 15 EBV‐negative BL lines were found to express C3 receptors. Furthermore, in vitro in association with EBV of six EBV‐negative cell lines resulted in the expression of C3 receptors in association with that of EBV‐determined nuclear antigen (EBNA). Rosette assays using erythrocytes coated with human C3b, C3bi, and C3d, inhibition of rosette formation with anti‐receptor antibodies, and flow cytometry analysis of stained cells demonstrated that EBV‐converted lines expressed C3b ami C3d receptors, CRl and CR2. Anti‐receptor antibodies recognized an average of40,700 anti‐CR1 and 140,000 anti‐CR2 binding sites on an EBV‐converted line(BL41/B95), whereas no specifie binding occurred on the corresponding EBV‐negative (BL41) cells, Because CR1 and CR2 are involved in B‐cell proliferation and/or differcntiation, enhanced expression of C3 receptors following the interaction between EBV and B celK and/or subsequent infection of the cells by EBV may provide a basis for positive control of B lymphocyte proliferation by EBV.
Induction of Proliferation and Differentiation of Leukaemic B Cells from Patients with Chronic Lymphocytic Leukaemia by Anti‐μ and Conditioned Medium
Authors: STEINBERG J. , MOORE M. A. S. , BERNHARDT B. , BONA C. A. , PLATSOUCAS C. D.
Published: June 2006

We investigated the growth and differentiation of leukaemic B cells from patients with B‐cell chronic lymphocytic leukaemia (CLL) in response to proliferation and differentiation factors present in conditioned medium and to anti‐immunoglobulin antibodies. Highly purified E‐rosette negative (E) B cells from 5 out of 15 patients with CLL exhibited moderate proliferative responses to Ml or 10 or 15 μ/ml of F(ab′)2 fragments of rabbit anti‐human μ‐chain specific antibody. Conditioned medium (CM), derived by stimulating human peripheral blood mononuclear leucocytes with PHA. induced significant proliferative responses of purified E cells in 13 out of 14 patients examined. The extent of these proliferative responses varied substantially, and was in the range of 2.6‐ to 91‐fold. Stimulation of purified L cells trom patients with CLL with both anti‐μ and CM resulted in significant proliferative in all l5 patients examined. These responses were significantly higher than thosc induced by CM alone (P<0.02). Synergism between CM and anti‐μ in inducing proliferative responses was observed in 11 out of 15 patients. Largely loukaemic B cell populations expressing on the cell surface more than one immunoglolobulin heavy chain isotype, exhibited significantly higher (P<0.009) proliferaiive response to CM and anti‐μ than those expressing IgM only. Highly purified E‐ peripheral blood or tonsil lymphocytes from all normal donors examined responded by proliferation to anti‐μ alone or to CM alone. Synergism in inducing prolifcrative responses was aiso observed when the cells were stimulated with both CM and anti‐μ. In addition to inducing proliferative responses, culture with CM of purified E‐rosctte negative, largely leukaemic, B cells from patients with CLL for 6 days at 37°C resulted in differentiation into immunoglobulin synthesizing and secreting cells. Synthesis and secretion of IgM were observed in 7 out of 10 patients examined. A switch to IgG production was observed in three patients. Morphological examination of E cells from patients wiih CLL after treatment wiih CM dcmonstrated thai these cells were differentiated into plasma‐like cells. These results suggest that leukaemic B cells from palicnls with CLL can be induced to proliferate and differentiate in response to growth and differentiation factors derived by mononuclear Icucocytes. in a manner similar to that of normal B cells.
Identification of Rat Erythrocyte Antigens with a New Non‐Radioactive Immunoprecipitation Technique
Authors: LAING P. , PARKAR B. A. , CULBERT E. J. , WATT G. J. , ELSON C. J.
Published: June 2006

In order to examine how rat erythrocytes stimulate eryihrroyte autoantibody production at the molceular level. we have identified rat erythrocyte antigens by immunoprecipition and western blotting using monoclonal antibodies and antisera A novel non radioactive immunoprecipitation technique was used, which employed biotin as a label and a luminescent detection system. The new method was validated by comparison with conventional immunoprecipitation using125I Glyeophorins of relative molecular mass (M1, 81,000 and 38,000 were found to be the major antigenic components of rat erythrocytes. while band 3(the most abundant erythrocyte membrane protein) was not retognized by rat‐specific antibodies. The same surface antigens were recognized by sera from mice producing erythrocyte autoantibodies and by sera from mice in which autoantibody production was suppressed. Nine other minor rat‐specific antifiens were identified by blotting. ranging in M1 from 23,000 to 147,000, Analysis of the integral membrane proteins of rat and mouse erythrocytes by sodium dodecyl sulphate (SDS) electrophoresis followed by silver or periodic acid‐Schiff (PAS) stains revealed differences beween the glyeophorins, but not between rat and mouse band 3. Thus, the major antigenic differences correspond to discernible biochemical differences between rat and mouse erthrocyte sialoglycuproteins.
Interferon‐Mediated Enhancement of Thyroid Major Histocompatibility Complex Antigen Expression
Authors: RAYNER D. C. , LYDYARD P. M. , DE ASSIS‐PAIVA H. J. , BIDEY S. , VAN DER MEIDE P. , VAREY A.M. , COOKE A.
Published: June 2006

Epithelial expression of class II antigens encoded by the major histocompaatility complex (MHC) has been proposed as a means by which autoimmune thyroid disease may be initiated and maintained. We studied a rat thyroid epithelial cell line (FRTL‐5), which constitutively expresses class I (OX18) but not class II (OX6orOX17)determinants to quantify in vitro MHC antigen induction using flow cytometry. Recombinant rat γ interferon (rlFN‐γ) induced dosedependent expression of OX6(l‐A) antigen at >48 h (maximum 80–90% of cells in culture at 100 U/ml). whieh was abrogated by DB‐I, a munoclonal antibody to rat IFN‐γ OXI7 antigen (I‐E) was also induced (86%) and OX18 (class I) markedly increased under these conditions Other thyroid‐active agents including the calcium ionophore A23187. dibutyryl cyclic AMP. thyroid‐stimulating autoantibodies from Graves’ disease patients (LATS), and TSH. caused no I‐A induction. Supernatants from spleen cells stimulaled with plant lectins(concanvialin A or phytohaemagglutinin). but not lectin alone. evoked substantial class II induction. which was inhibited yy DB‐1. These findings suggest that IFN‐γ is the central mediator of thyroid epithelial class II expression, FRTL'S provides a powerful model for the analysis of thyroid MHC elass 11 dynamies and a potential means of analysing the role of epithelial class II in autoimmune pathogenesis.
A Minority of Patients with Rheumatoid Arthritis Show a Dominant Rearrangement of T‐Cell Receptor β Chain Genes in Synovial Lymphocytes
Authors: SAVILL C. M. , DELVES P. J. , KIOUSSIS D. , WALKER P. , LYDYARD P. M. , COLACO B. , SHIPLEY M. , ROITT I. M.
Published: June 2006

The clonality of T lymphocytes isolated from the synovial fluid and peripheral blood of patients with rheumatoid arthritis was investigatcd by restriction enzyme fragment mapping of the rearrangement is of the β chain gene of the T‐cell antigen receptor. Three patients showed a dominant rearrangement amongst their synovial fluid T cells which was not seen in their peripheral blood T‐cell population, suggesting the prescnce of a predominating T‐cell clone. However, most of the patients examined (8 out of 11) demonstrated polyclonal T‐cell populations in both their synovial fluid and peripheral blood. Of four synovial fluid T‐cell lines investigated, one showed evidence of a dominant T‐cell clone.
Different Tc Response Profiles are Associated with Survival in the Murine Lymphocytic Choriomeningitis Virus Infection
Authors: THOMSEN A. R. , MARKER O. , PFAU C. J.
Published: June 2006

The pathogenicity of lymphocytic choriomeningitis virus (LCMV) inoculated intracercbrally (i.e.) varies with virus sirain and dose as well as with the mouse strain used as host. Recently, results have indicated that susceptibility to lethal disease correlates directly with the ability of the host to produce early and high vinis‐spccific Tc activity However, in the present studies we demonstrate that even though this holds true in many mouse/virus combinations, it does not apply in others Thus, in C3H mice infected with (moderately) high doses of Traub strain LCMV, early and high Tc activity was found despite a mortality rate of only 10–20%. Similarly. in C3H mice inoculated with the aggressive and docile suhstrains of UBC strain LCMV, which differ markedly in their pathogenicity for this mouse strain, similar kinetics of Tc induction were observed. Finally. in DBA/2 mice which do not die following infection with the otherwise lethal aggressive substrain. Tc induction could he found to be as efficient as m BALB/c mice, all of which die from acute LCM disease when infected with this virus isolate. The results indicate, therefore, that early and high Tc activiiy does not constitute a sufficient prerequisite for lethal disease. and that different Tc response profiles may he associated with low mortaility following i.e. inoculation with LCMV.
Selective Expansion of a CD3+CD4‐CD8‐ Subpopulation in Clinical Groups Associated with Human Immunodeficiency Virus Infection
Authors: MARCOS M. R. , GASPAR M. L. , DE LA HERA A. , TORIBIO M. L. , MÁRQUEZ C. , MILLÁN I. , MARTÍNEZ‐A C.
Published: June 2006

T lymphocytes (CD3+) without expression of CD4/CD8 surface antigens have recently been described in the thymus and peripheral tymphoid organs. We have conducted a retrospective analysis of the literature, seeking quantitative variations in this T‐cell subset in normal heterosexual controls, and in risk, pre‐AIDS, and AIDS groups, by means of the subtraction [CD3‐(CD4+CD8)] and the ratio 100×[CD3‐(CD4+CD8)]/CD3. Dramatic T lymphocytopaenia in AIDS patients and the progressive decay of CD4+ lymphocytes and increase of CD8+ lymphocytes throughout the clinical spectrum of HIV infection have been confirmed. Furthermore, we hereby demonstrate the selective expansion of CD3+CD4−CD8− lymphocytes, directly related to the clinical state in different clinical groups of infected people when compared with controls (P<0.05). The inverse relationship between the CD3+CD4−CD8− cell subset and other mature T‐cell subsets, mainly CD4+ (r=−0.49; P<0.01). suggests the existence of mutual regulatory interactions. These in vivo results, which are in agreement with those obtained in long‐term infected cultures, cannot be explained by direct cytopathic effects of the virus on the very few infected cells. Thus, the implication of the expansion of these functional precursors on the prognosis for infected people, and the paradoxes of the immunodeficiency, such as lymphoproliferation and autoimmune features, are discussed.
Generation of Reactive Oxygen Radicals by Human Phagocytic Cells Activated by Plasmodium falciparum
Authors: KHARAZMI A. , JEPSEN S. , ANDERSEN B. J.
Published: June 2006

The role of monocytes, macrophages and neutrophils in killing malaria parasites is well documented, and their involvement in malaria pathology has been suggested. However, the underlying mechanisms are not clear. The present study reports on the role of P. falciparum‐parasitized erythrocytes, free merozoites, and culture supernatant antigens in the generation of reactive oxygen radicals by human peripheral blood monocytes and neutrophils. Blood neutrophils and monocytes obtained from healthy individuals were isolated by density gradient separation. A human isolate of P. falciparum was grown in continuous culture. Parasitized erythrocytes and free merozoites were prepared from synchronized cultures. Soluble antigens from culture supernatants were purified by affinity chromatography using CNBr‐Sepharaose 4B columns bound to specific IgG. Oxidative burst response of neutrophils and monocytes were determined by oxygen consumption, superoxide production, and chemiluminescence. It was found that P. falciparum merozoites and the soluble antigens were capable of activating neutrophils and monocytes in vitro and resulting in the production of oxygen radicals by these cells. In conclusion, these findings demonstrate that malaria antigens are able to activate normal human blood phagocytes and result in generation of oxygen radicals by these cells. The released oxygen radicals can then contribute to both the destruction of the parasite and the pathology of malaria.
ELISA Assay for the Detection of a Dextran‐Binding Product Secreted by a T‐Cell Hybrid Th 1
Authors: PASANEN V. J. , MÖLLER G.
Published: June 2006

When CBA×BALB/c mice are immunized with Th 1 hybrid prepared by fusing spleen cells from Dx‐immunized mice with the AKR thymoma BW 5147, which secretes material that affects the anti‐Dx response in CBA mice, antibodies are produced that in the ELISA are shown to bind to the material secreted by Th 1 but not to the material secreted by parent BW 5147. A fraction of the Th 1 secreted material that starts eluting on DEAE chromatography in 20 mM Tris with 0.13 M NaCl is shown to bind lo Dx but not SRC or uncoated ELISA plates. This binding, like that of anti‐Dx Ig. can be inhibited with 1–10 μg/ml concentrations of free Dx. However, the affinity of the product of Th 1 for Dx is apparently lower than that of anti‐Dx Ig, since high concentrations of anti‐Dx interfere with the binding of Th 1 products to Dx. On gel permeation chromatography, a similar Dx binding material elutes in two molecular weight areas, a high area of the molecular weight of IgG and less and a low area of the molecular weight of 43,000 and above, and both are found in samples of Th 1 ascites and hyperimmune CBA anti‐Dx serum, but not (or less so) in normal CBA serum and not in the hyperimmune nude C57BI anti‐Dx serum. The Th 1‐derived material that affected the anti‐Dx response in previously shown experiments had similar elution characteristics. Since we also show that these fractions need not to contain Ig and that anti‐Th 1 minimally cross‐reacts with anti‐Dx IgM or IgG, it is likely that that fraction of Th 1‐secreted material represents a Dx‐binding, non‐Ig, anti‐Dx response‐regulating factor.
Cytotoxic T Lymphocytes: Blocked Differentiation at the ‘Poised’ Stage
Authors: GROMO G. , OCHOA A. C. , BACH F. H.
Published: June 2006

The effect of different concentrations of phorbol myristate acetate (PMA) on the development of cytotoxic cells was studied, PMA was selectively able to prevent the development of cytotoxic cells in a mixed leucocyte culture, while allowing the responding cells to proliferate. The higher concentration of PMA (10−5m) blocked both direct cytotoxicity and lytic activity in the presence ot lectin, while the lower concentration (10−8m) only prevented direct lytic function. The removal of PMA and subsequent addition of recombinant interleukin 2 (IL‐2) or IL‐2‐containing supernatants effectively reversed the effect of PMA with recovery of antigen‐specific lytic function of cells treated with 10−8m, while cells treated with 10−5m PMA only recovered lectin‐dependent cytotoxic ability.
Interaction of a Lymphokine with Normal Human Macrophages Results in Release of a Suppressor Factor for Mitogen‐Induced Immunoglobulin Synthesis
Authors: WARRINGTON R. J.
Published: June 2006

Normal human macrophage/monocyte cultures exposed to a suppressor factor produced by concanavalin A‐activated T cells (T‐SF), respond by releasing after 72 h a macrophage‐derived suppressor factor (Mφ‐SF). The Mφ‐SF inhibits pokeweed mitogen‐induced Ig synthesis but not T‐or B‐cell proliferation. Cycloheximide treatment of the macrophages does not interfere with generation of the Mφ‐SF, suggesting that de novo synthesis is not required. The factor is not preformed, for virgin maerophages do not contain Mφ‐SF. but it appears in macrophage cell lysates after exposure to T‐SF. The production of the Mφ‐SF is inhibited by the presence of 2‐mercaptoethanol. Both T‐SF and Mφ‐SF are l‐rhamnose inhibitable and the Mφ‐SF appears to be released as a high molecular weight complex which is dissociable into a low molecular weight form of a size similar to the T‐SF, i.e. ∼20,000. The T‐SF induced Mφ‐SF has some similarities with soluble immune response suppressor (SIRS) but differs from this factor in (a) its lack of effect upon lymphocyie proliferation (b) failure to induce conversion of T‐SF to Mφ‐SF by treatment with H2O2.
A Novel Arachidonic Acid Metabolite with Stimulatory Effect on PHA‐Induced Mitogenesis of Lymphocytes
Authors: JOHNSEN S. A. , GREEN K. , PAPADOGIANNAKIS N. , OLDING L. B.
Published: June 2006

The isolation of a novel arachidonic acid (Aa) metabolite from the supernatant of unstimulated human cord blood mononuclear leucocytes is reported. The metabolite, arbitrarily named ‘compound 4’is neither a known lipoxygenase nor a cyclooxygenase product,‘Compound 4’was added to PHA‐stimulated peripheral blood mononuclear leucocytes (PBML) from healthy blood donors, from mothers at term and from patients with immunodeficiency,‘Compound 4’induced an increase in the 3H‐TdR incorporation by the maternal PBML and by the PBML from patients with various immunodeficiencies such as Wiskott‐Aldrich syndrome and common variable immunodeficiency, whereas it had no effect on the proliferation of PBML from blood donors.
Thymus Localization of Monoclonal Antibodies Circumventing the Blood‐Thymus Barrier
Authors: STET R. J. M. , WAGENAAR‐HILBERS J. P. A. , NIEUWENHUIS P.
Published: June 2006

The binding in the rat thymus of mouse monoclonal antibodies which recognize determinants present on class II major histocompatibility complex antigens (HIS 19) and all T cells (HIS 17) was studied at several intervals after in vivo injection of a single dose (8 mg)of the monoclonal antibodies (MoAb). The MoAb, injected either intravenously or intraperitoneally. penetrated the thymus initially across the thymic capsule from the extravascular space. The extent to which the MoAb penetrated both cortex and medulla correlated with serum presence of the monoclonal antibody. These results suggest that the thymus cortex and medulla are permeable to MoAb present in the extravascular compartment. if this situation is a general phenomenon, which applies to circulating self non‐major histocompatibility complex antigens under physiological conditions, the current dogma relating to self‐tolerance needs to be reviewed.
The in Vitro B‐Cell Response to Pneumococcal Polysaccharides in Adults and Neonates
Authors: RUKERS G. T. , DOLLEKAMP E. G. , ZEGERS B. J. M.
Published: June 2006

In order to gain insight itito the relatively late appearance in ontogeny of responsiveness to T‐cell independent (TI) antigens, the in vitro human B‐cell response to type 4 pneumococcal polysaccharides (PS4, a human TI‐2 antigen) was studied. B cells obtained from adults differentiate into anti‐PS4 antibody forming cells upon culturing with PS4, but neonatal B cells obtained from cord blood fail to respond. The culture system used does, however, allow ihe differentiation of B cells reactive with TD antigens, for example ovalbumin. In order to evaluate the concept that in man the anti‐PS4 response is derived from a particular B‐cell subset we separated adult B cells on the basis of expression of the determinant recognized by the monoclonal antibody FMC7. The anti‐PS4 response is found mainly, bul not exclusively, in the FMC7+ subset. The selective unresponsiveness of neonatal B cells to TI‐2 antigens is not, however, due to the absence of FMC7+ B cells because, unlike adult blood B cells of which aboul 50% are FMC7+, 100% of neonatal B cells are FMC7+.
Dendritic Cells in Cutaneous, Lymph Node and Pulmonary Lesions of Sarcoidosis
Authors: MUNRO C. S. , CAMPBELL D. A. , DU BOIS R. M. , MITCHELL D. N. , COLE P. J. , POULTER L. W.
Published: June 2006

To determine whether dendritic cells (DC) are a consistent feature of lesions of sarcoidosis. we have used monoclonal antibodies to identify the HLA‐DR‐expressing populations of cells in cryostat sections of 15 lymph node, pulmonary and cutaneous lesions. The commonest HLA‐DR positive cells in granulomas were epithelioid and giant cells, although lymphocytes within granulomas and tissue macrophages around them were also positive. Dendritic eells with Langerhans cells (NA1/34+=OKT6+) and interdigitating cell (RFD1+) phenotype were consistency associated with granulomas only in skin lesions. In lymph nodes, interdigitating cells (NA1/34‐/RFD1 +/HLA‐DR++) were confined to paracortical zones as in normal nodes, although a small area of NA1/34+/RFD1+ cells was found in one of three nodes. In lung lesions NA1/34+/RFDI+ dendritie eells were uneommon or absent, except in one chronic case. We conclude that while sometimes present in extracutaneous sites. DC are not an essential feature of sarcoid lesions, and that cells of the classical macrophage group are the most signifieant HLA‐DR‐expressing population. We suggest that the presence ol DC in lesions of sarcoidosis may indicate an immunologieal response distinct from that causing granulomas to form. The variability of their involvement may have immunoregulatory significance.
Characterization of the Common Immunodominant Antigenic Determinant in the Lipopolysaceharide of Bacteroides fragilis by a Monoclonal Antibody
Authors: LINKO L. , WEINTRAUB A. , ARSTILA P. , PELLINIEMI L. J. , VILJANEN M. K.
Published: June 2006

The target determinant of a monoclonal antibody (MoAb) to Bacieroides fragilis lipopolysaccharide (LPS) was characterized by inhibition enzyme immunoassay (EIA), inimunoblotting (IB), immunofluorescence technique (IF) and electron immunocytochemical (EIC) technique. The MoAb has been shown to react positively with 96+ of B. fragilis isolates. LPS preparations from 14 different B. fragilis strains were tested by EIA and IB. Two LPS preparations did not react in any of the tests. In both preparations the o‐galactose was either lacking or present in low amount compared with the other LPSs. In addition, inhibition experiments with synthetic disaccharides confirmed that the target determinant is composed of β‐l, 6‐iinked galactose disaccharide. EIC showed that the target of the LPS‐MoAb is located on the surface of the outer membrane. These results show that the galactose chain present in LPS isolated from most B. fragilis strains contains the immunodominant antigenic determinant.
Production and Characterization of Human‐Human and Human‐Mouse Hybridomas Secreting Rh(D)‐Specific Monoclonal Antibodies
Authors: MACDONALD G. , PRIMROSE S. , BIGGINS K. , BOWMAN J. M. , BERCZI I. , FRIESEN A. D. , SEHON A. H.
Published: June 2006

Human monoclonal antibodies specific for the Rh(D) antigen were produced by cell lines generated by the fusion of pooled Epstein‐Barr virus (EBV)‐transformed B‐cell lines secreting Rh(D) antibodies with the murine myeloma cell line NS.l or with the human lymphoblastoid cell line HOA. 1. The selection of hybrids was achieved in RPMI 1640 medium containing HAT and ouabain. Higher fusion efficiency was obtained with the NS.l cell line; however the hybrids with HOA.l exhibited a greater elonal stability. The products of four clones (three human‐human and one human‐mouse) that consistently secreted antibodies for over 11 months were tested for specificity with a panel of red cells of various Rh phenotypes The supernatants of all four clones showed anti‐Rh(D) specificity but failed to react with the red cell Du phenotypes categorized as DV(Dw+) and DVI. Two of the three human‐human clones secreted IgM(λ) and the third lgG(x). The human‐mouse clone produced IgG(;v) antibody.
Selective Inhibition of Human Natural Killing and Antibody‐Dependent Cellular Cytotoxicity by a Polyanion
Authors: ROELLINGER S. , LOOS M. , LEIBOLD W.
Published: June 2006

A high molecular polyanion, Liquoid. was found to inhibit at nontoxic concentrations (12–50μg/ml) the natural killing (NK) and ihe antibody‐dependent cellular cytotoxic (ADCC) activity of human peripheral blood mononuclear cells selectively. Whereas NK of the K 562 target cell was slightly or not at ail affected, the spontaneous lysis of PDe‐B‐1, an EB V‐transformed B‐cell line, was strongly inhibited or even completely abolished. ADCC activity could only be inhibited by Liquoid if the target cells were mycoplasma‐free, while the polyanion had no effect when mycoplasma‐contaminated target cells were used. Liquoid did noi alter the target binding capacity of the NK effector cells and did not activate monocytes or induce other suppressive cells. Alpha interferon, but neither beta nor gamma interferon, was able to neutralize ihe NK reduction. These results suggest that Liquoid inhibits a target cell‐related, selective process in the post‐binding stage of NK cell lysis.
B Lymphocytes Respond Specifically to Phytohaemagglutinin after Liposome‐Dependent Transfer of Purified Phytohaemagglutinin Receptors
Authors: LETELLIER M. , DUPUIS G.
Published: June 2006

Phytohaemagglutinin (PHA) receptor glycoproteins purified by affinity chromatography from porcine splenic lymphocytes, were reconstituted into vesicles made of phosphatidylcholine and phosphatidylserine, concomitantly with Sendai virus fusogenic proteins HN and F. The vesicles were used as a vehicle to insert the PHA receptor glycoproteins into a highly enriched population of porcine B lymphocytes. Fluorescence analyses showed that 52±2% of the reconstituted B cells had incorporated the lectin receptors. The modified B lymphocytes were assayed for their response (tritiated thymidine incorporation into nucleic acids) to PHA, concanavalin A (Con A), or to lipopolysaccharide (LPS). The results showed that porcine B cells fused with vesicles containing only viral fusogenic proteins failed to respond to either PHA or Con A. Tritiated thymidine incorporation was similar to background values. The cells did, however, respond to LPS with values of label incorporation similar to those observed in the case of pre‐fused B lymphocytes. When purified B lymphocytes were fused with vesicles containing PHA receptors and viral fusogenic proteins, assays of thymidine incorporation showed a statistically significant (P<0.001) and specific response of the modified cells to PHA stimulation. Reconstituted cells cultured in the presence of PHA incorporated approximately nine times more radioactive label than pre‐fused cells or cells fused with vesicles containing only fusogenic viral proteins. In marked contrast, reconstituted B lymphocytes did not show any significant label incorporation above background level in response to Con A, but they retained their ability to respond to LPS. Our findings suggest that B lymphocytes can be made to respond specifically to PHA by insertion of appropriate lymphocyte‐derived receptors.
A Study of Cyclic Nucleotides as Second Messengers after Interleukin 2 Stimulation of Human T Lymphocytes
Authors: KNUDSEN T. E. , LARSEN C. S. , JOHNSEN H. E.
Published: June 2006

Interleukin 2 (IL‐2) was shown to induce a small but significant increase in the level of cGMP after 20 min stimulation and a subsequent fall afler 1 h in activated T lymphocytes. No change in the level of cAMP was observed. Addition of the cyclic nucleotide analogues dbcAMP or dbcGMP did not stimulate DNA synthesis. On the contrary, IL‐2‐induced [3H]thymidine incorporation was inhibited by these drugs. Further, the phosphodiesterase inhibitor theophyllin inhibited proliferation of activated T lymphocytes. Our results indicate that neither cAMP nor cGMP act as ‘second messengers’ for IL‐2 but support the theory that cAMP is a negative regulator of cell proliferation.
Monoclonal Antibodies against Murine Neonatal and Pregnancy‐Associated Natural Suppressor Cells Induce Resorption of the Fetus
Authors: GRÖNVIK K.‐O. , HOSKIN D. W. , MURGITA R. A.
Published: June 2006

Pregnant and neonatal/fetal mice have been shown to harbour naturally occurring inhibitory cells of both T and non‐T type. Non‐T suppressor cells present in the spleen of primiparous pregnant and newborn animals inhibit proliferative responses in autologous and allogeneic mixed lymphocyte reactions. Such cells can be positively selected for by agglutination with the B cell‐specific lectin soybean agglutinin (SBA). We generated rat IgG monoclonal antibodies against unique cell surface structures on the non‐T inhibitory cells, and cylotoxic pretreatment of spleen cells from pregnant or neonatal/felal mice largely abrogates their suppressive activity on proliferative responses. Furthermore, in vivo administration of such antibodies to pregnant inbred and outbred mice results in termination of the pregnancy or decreased litter size. Since injection of anti‐T cell IgG monoclonal antibodies does not interfere with the delivery of normal sized litters it is evident from these studies that the non‐T immunoregulatory cells, in contrast to T‐inhibitory cells, are of great importance in ensuring immunological homeostasis in the fetal‐placental environment during pregnancy in mice.
Amplification of T‐Cell Response to PPD by Epidermal Cell Suspensions Containing HLA‐DR‐Expressing Keratinocytes
Authors: TJERNLUND U. , SCHEYNIUS A.
Published: June 2006

The biological importance of the presence of class II transplantation antigens on highly differentiated epithelial cells such as keratinocytes in certain conditions, is still unknown. We have therefore investigated the antigen‐presenting capacity of separated human epidermal cells obtained from tuberculin‐reactive skin 6 days after intradermal injection of purified protein derivative (PPD). Earlier studies have shown a high percentage of HLA‐DR‐expressing keratinocyles at this time. Peripheral adherent blood cells were used as control stimulator cells a d highly purified peripheral blood T lymphocytes as responder cells. The T‐cell proliferation in response to PPD in the presence of autologous epidermal cells from normal and tuberculinreactive skin was measured by [3H]thymidine incorporation on day 6. The latter cell population, 76‐86% of which consisted of HLA‐DR‐expressing cells as judged by immunocytochemistry, induced a greater T‐cell response to PPD than do normal epidermal cells. This discrepancy in the T‐cell proliferation could not be explained by a difference in ihe numbers of anti‐Leu 6 or anti‐HLA‐DQ‐reactive Langerhans cells. The present data indicate that epidermal cell suspensions containing HLA‐DR‐expressing keratinocytes induce a greater T‐cell response to PPD than do normal epidermal cells.
Kinetics of Ca2+ Uptake into Lectin‐Induced Secondary Lymphocytes during Reactivation with Concanavalin A or Interleukin 2
Authors: WOLFF C. H. J.
Published: June 2006

The kinetics of Ca2+ uptake into lectin‐induced secondary lymphocytes was studied during reactivation to DNA synthesis with concanavalin A (Con A) or interleukin 2 (IL‐2). IL‐2 did not cause any significant uptake of Ca2+ into the lymphocytes, while Con A induced an accumulation of Ca2+ into the lymphocytes which reached a maximum 1 to 2h after the addition of the lectin. The time during which Ca2+ uptake occurred corresponded to the time of dependence on extracellular Ca2+ for lectin‐induced DNA synthesis. The increased rate of Ca2+ accumulation and the shortened Ca2+ dependence period of secondary lymphocytes as compared with primary lymphocytes could explain the ability of secondary lymphocytes to display an accelerated response, in terms of DNA synthesis, to re‐stimulus.
Role of Antigen‐Presenting Cells in the Cytotoxic T‐Cell Response to Minor Histocompatibility Antigens (MIHA)
Authors: PILARSKI L. M. , MOHAMED Z.
Published: June 2006

Although antigen‐presenting cells (APC) appear to be able to process minor histocompatibility antigens (MIHA) expressed on allogeneic cells and present them in association with intrinsic H‐2 of the APC in vivo, this does not occur in vitro. This could be due to fundamentally different mechanisms of antigen handling by APC in vitro, or it could represent compromised APC function. In order to distinguish these possibilities, we designed a system in which BALB/c spleen cells are transferred into an MIHA‐disparate irradiated host and allowed to reside for 2‐3 days; the spleen cells of the repopuiated host are removed and used as stimulator cells for the CTL response of primed BALB/c responder cells to DBA/2 MIHA. These cells are referred to as in vivo‐pulsed APC (IVP‐APC). Donor BALB/c cells are able to pick up DBA/2 MIHA after a passage in DBA/2 hosts and efficiently present MIHA to primed CTL precursors to generate DBA/2‐specific CTL. The donor cell type able lo pick up and present MIHA is present in spleen bul not thymus or bone marrow, is Thy‐1.2 negative, Ia+, and nylon wooladherent. Its stimulatory capacily is as efficient on a per cell basis as that of DBA/2 spleen cells. The generation of IVP‐APC requires repopulation of the irradiated host, which must express MIHA foreign to the BALB/c donor cells. When we attempled to generate IVP‐APC in H‐2 incompatible hosts, we found that, although the IVP‐APC could present H‐2 antigens, they were unable to present MIHA in association with intrinsic APC H‐2 antigens. Use of intra‐H‐2 recombinant strains as host mice indicated that compatibility of donor and host at the KI region of H‐2 was essential for the generation of IVP‐APC able to present apparently unprocessed MIHA to primed BALB/c responder cells. Thus, we were unable to reproduce the antigenprocessing function of APC encountering antigen in situ using an adoptive transfer method of pulsing APC with MIHA in vivo. In addition, we suggest that our results may impose constraints on the formulation of models to account for the association of MIHA with H‐2 antigens.
Characterization of Specific Immune Complexes in HIV‐Related Disorders
Authors: CARINl C. , MEZZAROMA I. , SCANO G. , D'AMELIO R. , MATRICARDI P. , AIUTI F.
Published: June 2006

Eighty‐seven seropositive subjects with HIV (human immunodeficiency virus) infection together with 20 normal controls with no history of any illness were investigated for the presence of circulating immune complexes (CIC) by the conglutinin binding assay (KgBA) and further studied for isotype characterization of CIC. Six out of 87 patients showing very high values for immune complexes (CIC) were studied for the presence of free antigen. In 3 out of 6 (1, IVcl; 1, III; I, IVa) we could detect by ultracentrifugation analysis the presence of specific HIV (p15) anti‐HIV (anti‐p15) and gp41‐anti‐gp41 CIC. Evidence in favour of this finding is supported by: (1) the presence of specific CIC (p15‐anti‐p15 or gp41‐anti‐gp41) seen only at pH 7.2; (2) the apparent presence of free antigen and specific HIV antibodies were only at pH 4.0. The relevance of this finding lies in the attempt to explain the occurrence of false seronegativity seen occasionally in symptomatic patients. Thus, the presence of CIC might perhaps interfere in the routine assay (i.e. ELISA) making the diagnosis difficult. All these considerations will have to be taken into account in the future handling of this disease.
Single Lipopolysaccharide‐Reactive B Cells in the Non‐Immune Mouse Spleen Cell Population Secrete Natural Multispecific Autoantibodies
Authors: RAGIMBEAU J. , AVRAMEAS S.
Published: June 2006

Starting from unimmunized BALB/c splenocytes, B‐cell clones were obtained by lipopolysaccharide (LPS) stimulation, and the frequencies of their anti‐TNP, anti‐BALB/c actin, and anti‐BALB/c tubulin secretion were determined. The culture conditions were then chosen so as to have one anti‐TNP precursor per positive well. Out of the 41 wells containing one anti‐TNP antibody‐secreting cell, nine (22%) also reacted either with actin or with tubulin and five (12%) with both actin and tubulin. Using horse red blood cells to which trinitrophenyl (TNP) had been coupled, spleen cell rosettes were prepared, enriched, micromanipulated, and cultured individually. Of the 500 micromanipulated and cultured TNP antigen‐binding cells, 28 were found to secrete antibodies directed against TNP. Eight of these 28 clones (28%) also reacted with either actin or tubulin, and five (17%) reacted with both actin and tubulin. The frequency of these multispecific clones is of the same order of magnitude as that obtained with the limiting dilution experiments. The results show that unimmunized BALB/c mouse spleen contains precursor cells that secrete natural multispecific autoantibodies.
IgD in Infant Saliva
Authors: GLEESON M. , CRIPPS A. W. , CLANCY R. L. , WLODARCZYK J. H. , HENSLEY M. J.
Published: June 2006

The ontogeny patterns of IgD in saliva were studied prospectively in 27 healthy full‐term infants. The concentrations and the proportion of samples with detectable levels of IgD were higher near birth. After 6 months of age IgD was rarely detected in saliva. There was an inverse assoeiation between the detection of IgD and IgA in saliva. There were no correlations between IgD and IgG or albumin. These observations support the theory that IgD synthesis and secretion at mucosal sites reflects a ‘physiological’ deficiency in IgA synthesis.
Enzymatic Fragmentation of an Unusual Human IgG2 (Kva) Myeloma Protein
Authors: VOGT R. A. L. , MICHAELSEN T. E.
Published: June 2006

The Kva IgG2(k) myeloma protein showed a complete resistance to papain in the presence of cysteine at neutral pH, and a higher resistance to trypsin and α‐chymotrypsin digestion than other IgG2 proteins. On the other hand, the Kva molecule was cleaved by pepsin at low pH to give the expected F(ab′)2 fragment. When the cleavage conditions were altered, it was possible to obtain Fab, Fc, and Fc′ fragments from this molecule as well. The Fab/c fragment and FacbFc′ complex were also obtained, which have not previously been reported from human IgG2 molecules. Incubation at elevated temperatures (45‐50°C) and/or lower pH resulted mainly in enzymatic attack on the C terminal side of the hinge. It was necessary to destroy the hinge by reduction or to expose the Kva molecule at 70°C or at lower pH (2.5) prior to digestion to facilitate enzyme digestion on the NH2 terminal side of the hinge. These results indicate that the hinge region of the Kva molecule has an unusually compact structure, which makes it extremely resistant to proteolysis
Patient‐Specific Heterogeneity of Antinuclear Antibodies as Revealed by an Isoelectric Focusing Immunoblot System
Authors: KOHLEISEN B. , KALIES I. , KÖLBLE K. , KALDEN J. R.
Published: June 2006

Ninety‐nine sera from patients with different rheumatic diseases (systemic lupus erythematosus, rheumatoid arthritis, progressive systemic sclerosis, and mixed and unidifferentiated connective tissue disease) were applied to a newly developed isoelectric focusing (IEF) immunoblot system for the demonstration of antinuclear antibodies. Nucleoproteins were separated according to their isoelectric points (pI) and immobilized onto nitrocellulose, and binding of serum antibodies was determined by an alkaline phosphatase labelled second antibody. 89.8% of all sera positive in indirect immunofluorescence assays with Hep 2 as substrate showed positive reactivity in IEF immunoblot. Furthermore, 88% of patients' sera negative on Hep 2 cells gave a positive reaction in IEF immunoblot. The predominant antibody banding pattern observed showed parallel bands in the acidic as well as the neutral pH ranges. Antibody specificities found in the IEF immunoblot system turned out to be patient‐specific, but no marker antibody for a discrete disease entity was obtained. Even when monoclonal antibodies or WHO standard sera were applied to nuclear antigen they exhibited heterogeneity in their binding pattern. Bands with the same pI were observed using sera from patients with different rheumatic disease entities. Immunodeletion experiments suggest the recognition of identical antigenic proteins by the different patients' sera.
Selective Modulation of the CD4 Molecular Complex by Pseudomonas aeruginosa Alkaline Protease and Elastase
Authors: PEDERSEN B. K. , KHARAZMI A. , THEANDER T. G. , ØDUM N. , ANDERSEN V. , BENDTZEN K.
Published: June 2006

The binding of monoclonal antibodies against CD4 was specifically inhibited by treatment of human CD4+ cells with either alkaline protease (AP) or elastase (Ela), purified from Pseudomonas aeruginosa. Binding of antibodies against CD3 (pan T), CD5 (pan T), CD8 (T suppressor/cytotoxic). HLA‐ABC, HLA‐DR, HLA‐DQ, HLA‐DP/DR, and β2 microglobulin was not inhibited by AP or Ela. Heat‐inactivation of the proteases at 65°C for 20 min or treatment with the metal chelator EDTA abolished the inhibitory activity of both proteases. These findings may serve to develop novel immunological methods for the isolation and study of the lymphocyte CD4 structure, which plays an important part in the immune response.
Meningeal Macrophages Reflect Lymphocytic Choriomeningitis Virus Pathogenic Phenotypes
Authors: WOODS S. J. , SARON M.‐F. , PFAU C. J.
Published: June 2006

Intracerehral (i.e.) intection of adult mice with lymphocytic choriomeningitis (LCM) virus can result in acute lethal central nervous system (CNS) disease which is the result of the host's thymus‐derived lymphocyte (T cell) response against the virus. Whether the specific effector function of the T cell is that of a cytotoxic cell (Tc) or a delayed‐type hypersensitivity cell (Ta) is still under debate. We assumed that if Td cells were important in pathogenesis then accessory cells in the brain (specificaily, glass‐adhcrent macrophages) might vary with the outcome of i.e. infection. We found that accumulation of macrophages in the brain (washed for meaninges and skull cap) appeared to be independent of the severity of the infection (controlled by the mouse strain as well as the strain and dose of virus used). However, differentiation of macrophages was clearly linked to whether or not the infection caused rapid death. In mice that were destined to survive, mycrophages became large, extensively vacuolated, and phagocytically active. In lethally‐infected mice macrophages were small and had poor phagocytic abilities, At present this dichntomy could be viewed as either a cause or a consequence of disease outcome. However. the data are not inconsistent with the hypothesis that Td lymphocytes may be of primary importance in pathogenesis.