bluuebunny/crossref_metadata_2025 huggingface dataset:
Infiltrating Neutrophils Differ from Circulating Neutrophils when Stimulated with C5a, NAP‐l/IL‐8, LTB4 and FMLP
Authors: MROWIETZ U. , SCHRÖDER J.‐M. , BRASCH J. , CHRISTOPHERS E.
Published: June 2006

In this study we report on functional characteristics of pustule as well as blood polymorphonuclear neutrophils (PMN) in patient suffering from relapsing bullous staphyloderma. Large numbers of viable PMN from newly formed pustules as well as from the peripheral blood were investigated. During the course of disease chemotactic migration, enzyme degranulation, superoxide‐anion generation and leukotriene B4 production were determined simultaneously.

The results revealed C5a‐ and NAP‐1/IL‐8‐specific dysfunction of pustule PMN as compared with blood PMN. In contrast, FMLP‐elicited functional activities of pustule PMN were only slightly affected.

Our findings provide evidence that in inflamed tissue invading PMN are regulated by in situ generated mediators. C5a produced by staph. aureus‐induced activation of the alternative pathway of the complement cascade represents predominant regulatory factor in situ. Furthermore, the results substantiate previous observations concerning different modulation of C5a and f‐met‐peptide receptors on human PMN.
Secretory Component is Bound to the Paraproteins in Sera of IgA and IgM Gammopathies
Authors: BOUVET J.‐P. , HOCINI H. , ISCAKI S.
Published: June 2006

Secretory immunoglobulins A (SIgM) and M (SIgM)were investigated in 20 sera containing high levels of monoclonal polymeric IgM or IgA. In the sera of patients suffering from Waldenström's macroglobulinemia (WM), the level of SIgA was found to be low, whereas that of SIgM was extremely high. Reciprocally, in the multiple myeloma (MM) patients, SIgA were increased and SIgM were dramatically decreased. Electrophoretic analysis showed these SIgA and SIgM to have the same monoclonal pattern as the corresponding paraprotein. Hence these molecules must originate from the malignant clone. The most likely mechanism involved is an intravascular formation of the secrelory‐like immunoglobulins. Free secretory component (SC) could diffuse passively from the digestive lumen and bind the circulating myeloma polymeric immunoglobulins. Such a possibility of in vivo binding of free SC to IgM and IgA polymers leads to a reconsideration of the secretory origin of SIgM and SIgA in normal human serum.
Generation of iC3 at the Interface between Blood and Gas
Authors: EKDAHL K. NILSSON , NILSSON B. , PEKNA M. , NILSSON U. R.
Published: June 2006

Earlier studies have shown that C3 can be denatured when blood comes in contact with a polystyrene surface. This study was undertaken to sec if similar denaturation of C3 occurs at the gas plasma interface which is found in all kinds of oxygenator used during cardio‐pulmonary operations. An in vitro system consisting of gas bubbling through human blood, serum or plasma was used. The generation of C3a, as an indicator of complement activation, and iC3 and iC3 fragments were monitored. Both C3a and iC3/iC3 fragments levels were increased during bubbling. In contrast to the C3a level, no reduction in iC3/iC3 fragments formation was seen in the presence of EDTA, indicating that il was independent of complement activation. The rate of iC3/iC3 fragments generation was unaffected by the composition of the gas (pure oxygen, pure nitrogen or air), suggesting that the denaturation of C3 indeed occurred at the serum gas interface. C3 and iC3/iC3 fragments were isolated from bubbled EDTA‐chelated serum by PEG precipitation and chromatography on FPLC, using a Mono S column and detected by two ELISAs, specific for native C3 and iC3/iC3 fragments. After 240 min approximately 20% of the total amount of C3 consisted of intact iC3 and it was confirmed that this population bound to human erythrocytes.
Glomerulopathy Induced by Graft‐Versus‐Host Reaction in the Rat. Requirement of Donor CD4+ T Lymphocytes and MHC Class II Incompatibility at the Lymphoid Compartment
Authors: ATEN J. , STET R. J. M. , WAGENAAR‐HILBERS J. P. A. , WEENING J. J. , FLEUREN G. J. , NIEUWENHUIS P.
Published: June 2006

Graft‐versus‐host reactions (GVHR) can he associated with several autoimmune phenomena involving the kidney as a target organ. By transferring lymphocytes of AO rats into complete Freund's adjuvant‐pretreated (AO × BN)F1 hybrids, a dose‐dependent GVHR with glomerulopathy was experimentally induced. IgM, IgG1, and IgG2a were deposited in the mesangial area and along the glomerular basement membrane. Fluted immunoglobulins from diseased kidneys hound to normal basement membranes and especially to laminin. Anti‐laminin reactivity was also present in sera from F1 recipients with GVHR. Parental CD4+ T lymphocytes were required and sufficient to induce GVHR and glomerulopathy in sublethally irradiated F1 hybrids. Using various F1 hybrids, MHC class II incompatibility was shown to be required for the induction of GVHR‐associated glomerulopathy. Across MHC class I incompatibility, GVHR without glomerulopathy could be induced, provided that both CD4+ and CD8+ donor T lymphocytes were administered. Finally, MHC incompatibility between donor T lymphocytes and the recipient non‐lymphoid compartment was found to be sufficient for the induction of GVHR, but not for GVHR‐associated glomerulopathy. The results indicate that alloreactive donor CD4+ T lymphocytes have to interact directly with MHC class II alloantigen bearing host B lymphocytes in order to stimulate the latter to produce (auto‐)antibodies. GVHR‐induced glomerulopathy shares several immunopathological features with HgCl2‐induced autoimmune glomerulopathy in the rat.
T‐Cell Receptor β‐Chain Gene Rearrangements of T‐Cell Populations Expanded from Multiple Sites of Synovial Tissue obtained from a Patient with Rheumatoid Arthritis
Authors: VAN LAAR J. M. , MILTENBURG A. M. M. , VERDONK M. J. A. , DAHA M. R. , DE VRIES R. R. P. , VAN DEN ELSEN P. J. , BREEDVELD F. C.
Published: June 2006

In this study T‐cell receptor (TcR) β‐chain gene rearrangements of T‐cell lines prepared from multiple sites (n=92) of synovial tissue derived from both knees of a patient with rheumatoid arthritis were analysed. In the majority of T‐cell lines, dominant TcR β‐chain gene rearrangements were detected, involving Cβ1 as well as Cβ2. The dominant rearrangement patterns of T‐cell lines from different tissue fragments showed significant variability, but some of the DNA restriction fragments were shared by T‐cell lines from multiple sites in both knees. The latter observation suggests that identical T‐cell clones may be present at different sites in the synovial tissue and in different joints. However, since many T‐cell lines yielded different rearrangement patterns, these data also indicate considerable heterogeneity of T cells in the joints. Apart from theoretical implications, this TcR heterogeneity of T cells within an individual patient also has practical consequences for studies on synovial T cells obtained by biopsy.
Prolonged Elevation of Intracellular Cyclic AMP Activates Interleukin‐1 Production in Human Peripheral Blood Monocytes
Authors: SERKKOLA E. , HURME M. , PALKAMA T.
Published: June 2006

The capability of elevated intracellular cyclic AM P concentration to activate IL‐l gene expression and protein production was examined in human peripheral blood monocytes. In accordance with previous studies it was observed that the transiently elevated cyclic AMP (induced either with prostaglandin E2 or with the direct adenylate cyclase activator, forskolin) was not a sufficient signal to activate IL‐l production. However, if the degradation of cyclic AMP was inhibited with isobutyl‐methyl‐xanthine (IBMX). IL‐l production was strongly activated. This prostaglandin E2 plus IBMX effect could also be mimicked with high concentrations of the cell permeant structural cyclic AMP analogue, dibutyryl cyclic AMP. The cyclic AMP4nduced IL‐l production differed in some aspects from the bacterial lipopolysaccharide‐induced IL‐l production: (1) the kinetics of both IL‐l gene expression and protein production was much slower: (2) the IL‐1β gene expression was superinducible by inhibiting the protein synthesis with cycloheximide. Thus these data suggest that prolonged elevation of cyclic AMP is alone a sufficient signal to activate IL‐l production.
Antibodies to MHC Class II Peptides are Present in HIV‐1‐Positive Sera
Authors: ZAITSEVA M. B. , MOSHNIKOV S. A. , KOZHICH A. T. , FROLOVA H. A. , MAKAROVA O. D. , PAVLIKOV S. P. , SIDOROVICH I. G. , BRONDZ B. B.
Published: June 2006

Seventy‐five per cent of sera from HIV‐I‐infected individuals bind to the human B‐lymphoma cells bearing the major histocompatibility class II molecule in enzyme‐linked immunosorbent assay (ELISA). The binding is caused by the antibodies against the class II molecule present in the serum samples which prevent the interaction of murine anti‐HLA.DR monoclonal antibody with B lymphoma in FACS analysis. The three highly conserved amino acid sequences in α‐ and β‐ chains of the class II molecule and three homologous fragments in HIV‐1 gp120 and gp41 were identified by computer search and synthesized. Using these peptides it was demonstrated that 28–48% of HIV‐positive sera contain antibodies that cross‐react with the peptide of HIV‐I origin and with the peptide from the class II molecule as well.
Molecular Events in Late Stages of T‐Cell Functional Maturation
Authors: NELSON P. J. , GELLER R. L. , PODACK E. , BACH F. H.
Published: June 2006

Peripheral blood lymphocytes activated with either the calcium ionophore A23187 or the combination of anti‐CD2 monoclonal antibodies. 9.6+VIT13, undergo blast formation and proliferation but do not develop cytolytic activity. These proliferating blasts, referred lo as pre‐effector blasts because they do not yet express cytolytic function, respond to stimulation with interleukin‐2 (IL‐2) by further proliferation and development of cytolytic activity, i.e. they become effector cells. Pre‐effector blasts activated with 9.6 +VIT13, but not A23187‐activated pre‐effector blasts, also respond to stimulation with interferon‐gamma (H N‐γ) by becoming cytolytic effector cells. This report examines gene expression (by Northern blot analysis) in pre‐effector blasts and during the transition from the pre‐effector to the effector stage. The data presented here provide further support for the concept that A23187 activation drives T cells to become dividing blasts that are appropriately referred to as ‘pre‐affector’ cells in that these blasts do not express transcripts for granzyme A or perforin mRNA but are driven by IL‐2 to do so in parallel with the acquisition of cytotoxic function. Cells are apparently driven by 9.6 +V IT 13 to a later stage of functional maturation than by A23187 activation; 9.6 +VIT13‐activated pre‐effector blasts express mRNA for both granzyme A and perforin, even though these blasts do not express cytolytic activity. Activation via A23187 results in lower expression of the proto‐oncogene c‐myb relative to that found in either 9.6‐VIT13 or OKT3‐activated cells.
Role of Cross‐Linking in Stepwise Activation of T Cells
Authors: GELLER R. L.
Published: June 2006

Activation of T cells is a complex process which we have hypothesized involves a series of functional intermediates possessing some, but not all, of the characteristics of fully functional effector cells. We have identified two such functional intermediates, the poised T cell (po Tc) and the pre‐effector T cell (pe Tc). poTc do not proliferate or mediate cytolytic activity but are responsive to help in the form of interleukin 2 (IL‐2); peTc are proliferating cells which arc not cytolytic. Here the role of T‐cell receptor/CD3 complex cross‐linking in generating these functional intermediates is examined and it is shown that cells can be driven to different stages of functional maturation depending upon the sequence of antibody binding to CD3 and cross‐linking. Highly purified T cells can be activated to poTc stage if they are first labelled with the anti‐CD3 monoclonal antibody OKT3 and then cross‐linked with goat anti‐mouse IgG‐coated beads. If the OKT3 antibody is first bound to the IgG‐coaled beads and then added to highly purified T cells, peTc are generated. In both cases the intermediates can be driven to become fully functional effector cells by the addition of IL‐2. Finally, by removing the OKT3‐bound beads during the activation process, were are able to show that po Tc and pe Tc are sequential intermediates along the same pathway of activation.
Role of ELAM‐1 in Adhesion of Monocytes to Activated Human Endothelial Cells
Authors: LEEUWENBERG J. F. M. , JEUNHOMME T. M. A. A. , BUURMAN W. A.
Published: June 2006

The role of ELAM‐1 in the adhesion of monocytes to HUVEC, activated for 4 h with TNF, was studied using MoAb ENA2 directed against ELAM‐1. In a standard adhesion assay at 37 C, F(ab')2 fragments of f ENA2 did not, or weakly inhibited adhesion. When metabolic activity of the monocytes was reduced by (i)fixing the monocytes, (ii) performing the adhesion assay at 4°C, and (iii) combining the forementioned conditions, the adhesion of the monocytes was strongly blocked by ENA2 and less effective or not by MoAb IB4 anti‐CD18. The pattern of adhesion of monocytes to HUVEC, activated with TNF assessed at 4°C. paralleled ELAM‐1 expression on the endothelial cells. Maximal inhibitory effect of ENA2 on adhesion was shown 5 h after activation of HUVEC, at which ELAM‐1 expression was also maximal. Adhesion assessed at 37°C remained enhanced for at least 24 h, whereas the inhibitory effect of ENA2 followed ELAM‐1 expression. Specific involvement of ELAM‐1 was also confirmed using ELAM‐1 transfected COS cells. These results indicated that monocytes express a counter structure for ELAM‐1 and that this counter structure is involved in adhesion.
A Study of Donor Alloreactivity, which may predict Acute Graft‐Versus‐Host Disease in HLA Identical Bone Marrow Transplantations for Early Leukaemia
Authors: JOHNSEN H. E. , BOSTROM L. , MOLLER J. , JORGENSEN J. A. , JENSEN L. , RINGDEN O.
Published: June 2006

Recent studies performed in Seattle, USA have suggested that pretransplant assignment of high or low donor alloreactivity may predict acute graft‐versus‐host disease (aGvHD) after allogeneic HLA identical marrow transplantation for acute leukaemia. The effect of such pretrunsplant assignment was studied in a Scandinavian population of 114 consecutive transplantations for acute and chronic leukaemias in 1st remission (n= 74) or chronic phase (n= 40) performed between 1975 and 1989.

The selected cut‐off value for discriminating between donors of high and low responding capacity (DRC) was based on distribution plots of results from the pretransplant mixed lymphocyte culture (MLC) and chosen as the median value (80% normalized response). Then 57 donors were assigned with high DRC and 57 donors assigned with low DRC. Kaplan‐Meier estimates of the probability of patients to develop Grade M or higher aGvHD in receipt of high or low responder donor transplants were compared by univariant analysis. The patients in first remission or chronic phase transplanted with bone marrow from donors assigned as high or low responders had a 36.1% and 10.6% risk for aGvHD, respectively, a difference found to be significant by log rank test (chi‐squared = 10.1, d.f.= 1, P= 0.0015).

Subsequent studies of the cellular and humoral requirements for this predictive response of donor cells, by blocking with cytokine specific antibodies, addition of excess of recombinant human cytokines and scanning of lymphocyte subsets during the response, showed that the response against pool cells mostly depended upon IL‐2 responding cells with the phenotype CD3+, CD4+, CD8‐, CD25+, CD16‐.

It is concluded that prospective studies of alloreactivity as a risk factor should be performed to confirm the above findings,
The Direct Binding of an HIV Fragment with Granulocytes from Healthy Subjects and Infected Patients
Authors: GABRILOVICH D. I. , KOZICH A. T. , MOSHNIKOV S. A. , POKROVSKY V. V.
Published: June 2006

The binding of the synthetic fragment from the CD4 binding site of HIV (gp120) with polymorphonuclear neutrophils (PMN) in 13 healthy donors and 31 HIV‐infected patients was studied using a biotin‐streptavidin‐texas‐red complex. The largest percentage of PMN‐bound peptide was reached at a final peptide concentration of 10 μg/ml. The increase of peptide concentration did not raise the per cent of positive PMN. Preliminary incubation of PMN or mononuclear cells with non‐biotinylated peptide abolished subsequent binding of these cells wish biotinylated peptide, while preliminary treatment of the cells by anti‐CD4 monoclonal antibody did not lead to such abrogation. It was revealed that about 14% of PMN but no lymphocytes from healthy donors were able to bind peptide, The number of such PMN in HIV‐infected patients was significantly less (6.3±8.9%, P<0.05). A connection between peptide‐bound PMN and their functional activity was found. The percentage of such cells was 13.0±11.5% in patients with normal values in a stimulated nitroblue tetrazolium reduction test and only 2.6±2.8% in patients with low values in this test (P<0.05).
A Supportive Role of Neurol Cell Adhesion Molecule (NCAM) in Adhesion Between Leukaemic Blasts and Cytotoxic Lymphocytes
Authors: PALUCKA A. K. , PORWIT A. , REIZENSTEIN P.
Published: June 2006

The expression and function of neural cell adhesion molecule (NCAM, CD56, Leu 19) on leukaemic blasts was investigated. The expression of NCAM was frequent (64%) in 14 studied cases of acute myeloid leukaemia (AML) but not in chronic lymphocytic leukaemia (CLL: 1/3 cases positive) or immunocytoma (IC; no positively). No correlation with the expression of other AM (intercellular adhesion molecule‐1 (ICAM‐1), leucocyte function antigen‐1 (LFA‐1), VLA beta chain) or with AML type, according to FAB classification, was observed. Blocking of NCAM with anti‐Leu 19 MoAb on AML targets resulted in a significant decrease of their susceptibility to LAK killing and in inhibition of conjugate formation. In the case of B prolymphocytic leukaemia (B‐PLL) which did not express ICAM‐1 or LFA‐1 but was NCAM+, a complete resistance to LAK activity and lack of conjugate formation was observed. Blocking of NCAM on LAK effectors did not decrease their cytotoxic activity. Our results suggest that NCAM, in the presence of other AM, may have a supportive role in adhesion of leukaemic targets to LAK effectors.
Preferential Interaction of the CD4+29+/45RA‐ Subset of Human CD4+ T Lymphocytes with an Antibody against the Cell‐Membrane Ganglioside GD3
Authors: WASIK M. A.
Published: June 2006

This study shows that the two major subpopulations of CD4+ T lymphoyctes, defined on the basis of differential expression of the CD29 and CD45RA antigens, show significant differences in reactivity with a monoclonal antibody against the GD3 ganglioside. Double staining studies showed that the GD3 ganglioside is predominantly expressed on cells from the CD4+29+ 45RA‐ subsets. The preferential interaction of the CD4+ 29+/45RA‐ cell subset with the anti‐GD3 MoAb was further confirmed by the proliferative and calcium‐flux studies. Accordingly, the reciprocal, CD4+ (29‐)/45RA+ subset was unable to proliferate in response to the anti‐GD3 MoAb alone. Although it did show a significant mobilization of calcium ions and proliferation to IL‐2 when stimulated with the anti‐GD3 MoAb, these responses were much less pronounced than the responses of the CD4+ 29+/45RA‐ subset. Finally, when two T‐cell stimulating monokines, IL‐1 and IL‐6, were tested for the ability to modulate the anti‐GD3 mediated proliferation, only the former, but not the latter was able to enhance the proliferation. Although the natural ligand for the GD3 ganglioside remains unknown, the data presented here provide further evidence in support of the notion that the T‐cell surface molecules different from the T‐cell receptor MHC antigen complex may contribute to the preferential activation of one of the CD4+ T lymphocyte subsets.
Human Umbilical Vein Endothelial Cells Synthesize S‐Protein (Vitronectin) In Vitro
Authors: BERGE V. , JOHNSON H. , HØGÅSEN K. , HETLAND G.
Published: June 2006

S‐protein, also named vitronectin, is a multifunctional glycoprotein with molecular weight (MW) of about 75 kDa and a serum concentration of 0.14‐0.60 mg/ml. It is synthesized mainly in the liver, but synthesis has also been found in monocytes/macrophages. We used human umbilical vein endothelial cells (EC) which were incubated wilh agarose beads, an activator of the alternative complement pathway. By radioimmunoassay (RIA) based on monoclonal and polyclonal S‐protein antibodies, we delected S‐protein on harvested agarose beads. The time dependent increase m the amount of S‐protein was significantly reduced by the presence of cycloheximide (10 μg/ml) in the cell cultures. We also found a strong binding of S‐protein antibodies to agarose beads preincubated in native serum, which was strongly reduced (70‐80%) by inactivation of the alternative complement pathway (50°C, 20 min). Our results show that EC synthesize S‐protein in vitro.
Purification, Subunit Characterization and Ultrastructure of Three Soluble Bovine Lectins: Conglutinin, Mannose‐Binding Protein and the Pentraxin Serum Amyloid P‐Component
Authors: ANDERSEN O. , FRIIS P. , NIELSEN E. HOLM , VILSGAARD K. , LESLIE R. G. Q. , SVEHAG S. E.
Published: June 2006

Conglutinin and mannose‐binding protein (MBP) are members of the C‐type lectins which are widely present in mammalian plasma. Serum amyloid P‐component (SAP) is a member of the pentraxin family with lectin properties. A scheme for the partial purification of all three lectins by carbohydrate affinity chromatography and selective elution was developed. The purification was monitored by SDS‐PAGE, Western blotting and electron microscopy. Binding of the lectins to Sephadex‐iC3b. their collagenase sensitivity, and the size and antibody reactivity of their subunits was investigated. The demonstration, by SDS‐PAGE, of 25‐kDa subunits, which were unaffected by collagenase treatment but bound to Sephadex‐iC3b and antibodies to human SAP, indicated the existence of bovine SAP. Bovine conglutinin (BK) also showed calcium‐dependent binding to Sephadex‐iC3b, whereas bovine MBP did not. The binding of BK was inhibitable with GlcNAc. A 3000‐fold increase in BK activity (ELISA) was obtained in eluates from Sephadex iC3b. SDS‐PAGE analyses of BK and MBP revealed subunits with an Mr of 43 kDa and 30 kDa, respectively. These subunits were sensitive to collagenase treatment which reduced the Mr to 20 kDa. Electron micrographs revealed a prominent flexible tertramer molecule (diameter 96 nm) in the BK preparations, a predominantly hexameric structure (diameter 30 nm) in the MBP preparations, and single annular pentameric disc‐like molecules (diameter 11 nm) in the SAP preparations.
From Pathology to Physiology of the Human T‐Lymphocyte Receptor
Authors: REGEIRO J. R. , TIMÓN M. , PÉREZ‐ACIEGO P. , CORELL A. , MARTÍN‐VILLA J. M. , RODRÍGUEZ‐GALLEGO C. , GÓNGORA R. , ARNAIZ‐VILLENA A.
Published: June 2006

The recent description of a selective human CD3γ deficiency and other T‐cell receptor (TCR)/CD3 structural and functional defects, together with previous biochemical data on the structure and interactions of the TCR/CD3 complex, may aid in elucidating the physiology of this multi‐subunit membrane ensemble. CD3γ seemed to be required for the commitment and thymic maturation of an important fraction of T lymphocytes to the CD8 (but not CD4) lineage, perhaps by participating with the CD8 co‐receptor in the instructive signal delivered throtigh the αβ TCR during intrathymic positive selection by HLA class I molecules. The homologous CD3δ component would, in contrast, be necessary for the selection of CD4 lymphocytes by HLA class II molecules. The interaction of CD4 and CD8 with the TCR/CD3 complex during antigen recognition may thus be asymmetrical, taking place through CD3δ and γ respectively. Also, the existence of in vivo functional TCR/CD3 hemireceptors (lacking either CD3γ or CD3δ is suggested, and defects in their relative amount on the T‐cell surface may disrupt unresponsiveness to self antigens and generate autoimmunity.
Involvement of Interleukin‐2 and Interferon‐Gamma in the Immune Response Induced by Influenza Virus Iscoms
Authors: VILLACRES‐ERIKSSON M. , BERGSTRÖM‐MOLLAOGLU M. , KÅBERG H. , MOREIN B.
Published: June 2006

Splenocytes from mice primed with influenza virus envelope proteins incorporated in iscoms, as micelles or as infectious virus, were restimulaled in vitro with the same antigen. Interleukin‐2 (IL‐2) and interferon‐gamma (IKN‐γ) were assayed in the supernatants of such cultures. Influenza virus iscoms induced IL‐2 and IFN‐γ responses in restimulation experiments that were antigen specific and significantly higher than those induced by micelles or infectious virus. Serum samples collected at the end of the experiments were analysed for the antibody response and profile. The antibody titres induced by iscoms were of a similar order of magnitude as those induced by infectious virus, and were about 18 times higher than the titres induced by micelles. In mice immunized with iscoms or infectious virus the most abundant antibodies were of the IgG1 and IgG2a, isotype, and the IgE response was low. We conclude that immunization with iscoms stimulates the Th1‐like subtype of murine T lymphocytes.
Alteration in T Cell/Macrophage Ratio may Reveal Lymphocyte Proliferation Specific for the Triggering Antigen in Reactive Arthritis
Authors: SIEPER J. , BRAUN J. , WU P. , KINGSLEY G.
Published: June 2006

It has previously been shown that synovial fluid (SF) mononuclear cells (MNC) from patients with reactive arthritis (ReA) and some patients with undifferentiated oligoarthritis (UOA) respond specifically to the triggering bacterium (specific responders). However, in some patients there is a response to two or more bacteria (non‐specific responders) and in a third group no response is found (non‐responders). We assessed whether the proportion of synovial MNC which were macrophage‐monocyte (MaMo) differed among the specific responder, non‐specific responder and non‐responder groups. There was no difference between the specific (33±9) and the non‐specific(32±26)groups; non‐responders had a higher percentage of MaMo (61.3±31%) although the difference was not significant. We also investigated whether the specificity of the response to antigen in ReA or UOA SF was altered by changing the T‐cell/MaMo ratio. In all five specific responders the immune response remained specific whatever the ratio tested. However, four of the five non‐specific responders, but none of the non‐responders, developed a specific response to one of the tested antigens by increasing the T cell/MaMo ratio. We conclude that in some patients with a non‐specific response, alteration of the T cell/MaMo ratio uncovers a specific response which may identify the triggering antigen.
T‐Cell Epitopes on the Human Acetylcholine Receptor α‐Subunit Residues 10‐84 in Myasthenia Gravis
Authors: ÅHLBERG R. , YI Q. , ENG H. , PIRSKANEN R. , LEFVERT A. K.
Published: June 2006

In myasthenia gravis the production of anti‐acetylcholine receptor antibodies is modulated by acetylcholine receptor‐specific T cells. Most B‐and T‐cell epitopes are located on the α‐subunit of the receptor. In order to map the fine specificity of the antigen‐specific T cells in myasthenia gravis, T‐cell stimulation in response to 70 hexapeptides was studied in 24 patients and 24 healthy individuals. The hexapeptides overlapped with one amino acid and represented residues 10‐84 of the NH2‐terminal part of the α‐subunit of the receptor. The IFN‐γ secretion from single T cells was used to detect T‐cell stimulation.

A significanl difference in the T‐cell response to several of the peptides was found between patients and healthy controls. The majority of the hexapeptides induced T‐cell stimulation in at least one of the patients. Peptidc‐induced T‐cell stimulation was evident in all but one of the patients. The results indicate that different epitopes and multiple T‐cell clones are involved in the T‐cell recognition of the acetylcholine receptor.
Production of a Suppressor of Lymphocyte Proliferation by Two Human Oral Carcinoma Cell Lines
Authors: RICE S. Q. J. , CRANE I. J. , SCULLY C. , PRIME S. S.
Published: June 2006

This study examined the production of an immunosuppressive factor by the KB and H191 human oral squamous carcinoma cell lines. Conditioned media (CM) from both cell lines markedly inhibited mitogen‐ and alloantigen‐induced proliferation of normal human and rat peripheral blood lymphocytes. By contrast, the proliferation of an exponentially‐growing fibroblast cell line remained unchanged by CM. The immunosuppressive factor appeared to act after lymphocyte commitment as indicated by continued blast cell formalion, the failure of CM to suppress resting lymphocytes and the fact that CM caused maximum inhibition of lymphocyte proliferation 72 h after the addition of PHA. The addition of exogenous IL‐2 did not counteract lymphocyte suppression. Inclusion of indomethacin and isoniazid during cell culture did not significantly alter the degree of suppressive activity. Mycoplasma contamination was absent and CM did not act directly with the thymidine or mitogen. The factor was heat stable at 50 C, acid labile and had a molecular weight in execss of 300 kDa. The results demonstrate that human oral squamous carcinoma cell lines produce an immunosuppressive factor that may have a role in tumour evasion of the host immune response.
Unusual Sequence of Immunoglobulin L‐Chain Rearrangements in a Gamma Heavy Chain Disease Patient
Authors: HAUKE G. , SCHILTZ E. , BROSS K. J. , HOLLMANN A. , PETER H. H. , KRAWINKEL U.
Published: June 2006

Patients with gamma heavy chain disease (γ‐HCD) generally produce incomplete immunoglobulin (Ig) γ‐heavy chains (yγ‐HCD protein) which cannot associate with light chains (IgL). In most patients Bence Jones proteins (BJP) are not observed. However, in the 61‐year‐old patienl WIN we found γl‐HCD proteins and λ BJP in serum and urine. WIN ‐γl‐HCD protein does not carry the Ig Fd region, has a molecular weight of 33.5 kDa, and the seven N‐terminal amino acid residues are not translated from any of the known immunoglobulin heavy chain (IgH) gene sequences. These residues are followed by the Cγl‐hinge region. In DNA from peripheral blood lymphocytes of patient WIN we found bands representing dominant rearrangements in one ofthe two alleles of the IgH, IgK and Igλ locus. Taken together, the data from protein and DNA analysis strongly suggest, albeit do not formally prove, that one dominant B‐cell clone which carries it rearranged and a non‐rearranged allele of each Ig locus produces γ‐HCD protein and λ BJP. The productive λ‐gene rearrangement in this clone thus has not been preceded by abortive rearrangements in both γ‐locus alleles. Lymphocytes with an unusual sequence of IgL‐chain gene activation seem to be involved in the case of γ‐HCD described here.
Staphylococcus albus‐Induced Protein Kinase C Translocation in Human Neutrophils: The Effect of Opsonization, Cytochalasin B, Pertussis Toxin, Intra‐ and Extracellular Calcium, and R59022
Authors: OBEL N.
Published: June 2006

Membrane‐associated protein kinase C has been p[roposed to tbe essential in transmembrane signalling systems activating neutrophils. A main function of the neutrophil is phagocytosis and killing of microorganisms. Nevertheless, previously published reports mainly have described the effect of artificial or soluble stimulators upon neutrophil protein kinase C activity. Therefore membrane‐associated protein kinase C was studied in neutrophils stimulated by Staphylococceualbus. The bacteria were found to induce a striking increase in membrane associated protein kinase C, an effect which depended upon a previous opsonization of the bacteria. Cytochalasin B which inhibits phagocytosis, was shown to abrogate S. albus‐induced protein kinase C translocation. Chelation of intracellular calcium totally abolished S. albus‐induced protein kinases C translocation, a phenomenon that could not exclusively be ascribed to chelation of extracellularcalcium. The diacylglycerol kinase inhibitor R5902, which has been reported to increases endogeneous diacylglycerol accumulation, nearly doubled the effect of S. albus upon membrane associaed protein kinase C. Pertussis toxin in concentrations which completely inhibited fLMP‐induced superoxide generation did not affect S. albus‐induced protein kinase C translocation. It is the cell membrane, a phenomenon that reies upon enhanced diacylglycerol production and calcium transients and occurs independently of pertussis toxin‐inhibitable G‐proteins.
Antibodies to Eukaryotic, including Autologous, Native DNA are Produced during BK Virus Infection, but not after Immunization with Non‐Infectious BK DNA
Authors: REKVIG O. P. , FREDRIKSEN K. , BRANNSETHER B. , MOENS U. , SUNDSFJORD A. , TRAAVIK T.
Published: June 2006

The contemporary view concerning the origin of anti‐dsDNA antibodies is that eukaryotic dsDNA is not immunogenic. Results presented here, however, show (1) that inoculation of rabbits with BK virus elicits antibodies to eukaryotic, including autologous, dsDNA. (2) that the transition from a non‐immunogenic to an immunogenic state of autologous dsDNA depends on productive infection with BK virus, and (3) that inoculation with protein‐free circular BK dsDNA initiates both infection in vivo and production of antibodies to autologous dsDNA. Non‐infectious linearized BK dsDNA did not elicit any anti‐dsDNA antibodies, while the same DNA molecule, when complexed with methylated bovine serum albumin, elicited anti‐dsDNA antibodies solely recognizing BK dsDNA. Neither of the two linearized BK dsDNA preparations initiated infection. Using two different techniques, we could demonstrate that two separate sets of anti‐dsDNA antibodies were produced during viral infection; one recognizing BK dsDNA, and the other recognizing autologous dsDNA. Thus, in contrast to previous assumptions, autologous dsDNA may be immunogenic. Based on the present results, we propose that autologous dsDNA can be rendered immunogenic through complex formation with viral DNA binding protein(s) such as the structural protein VP1 or the tumour antigen T. Such DNA protein complexes may bypass a pulative T‐cell tolerance to autologous dsDNA.
The Effect of Calcium Mobilization on LPS‐Induced IL‐1β Production Depends on the Differentiation Stage of the Monocytes/Macrophages
Authors: HURME M. , VIHERLUOTO J. , NORDSTRÖM T.
Published: June 2006

The role of elevated intracellular Calcium concentration [Ca2+]i in the LPS‐induced activation of interleukin‐1β (IL‐1β) production was examined in cells representing different stages of myeloid differentiation (undifferentiated monocytic leukaemia cell line THP‐1, THP‐1 cells induced to adherent, macrophage‐like cells by phorbol ester treatment and normal peripheral blood‐derived adherent monocytes). LPS did not elevate the [Ca2+]1, as measured by the Fura‐2 fluorescence technique. When these cells were stimulated with LPS in the presence of the calcium ionophore A23187, a clear increase in the IL‐ 1β protein production was observed in the undifferentiated THP‐1 cells but not in the more differentiated cell types. This ionophore‐induced increase was also seen in the IL‐1β mRNA levels. Thus these data confirm the previous findings demonstrating that elevation of (Ca‐2+]1 is not involved in the LPS‐dependent signal transmission. However, the LPS‐induced signals are greatly potentiated by the elevated [Ca2+]i, but only in undifferentiated monocytic cells.
Resistance to Collagen‐Induced Arthritis in Biozzi Mice is Not Associated With a Defect in Autoantibodies to Accessible Epitopes on Cartilage Collagen
Authors: BOUVET J.‐P. , FRANC B. , ZYAD A. , COUDERC J.
Published: June 2006

In order to understand why most strains of Biozzi mice are unexpectedly either arthritis‐resistant, or susceptible, we investigate the autoantibody reactivity against matrix‐embedded collagen molecules. We show a dose correlation between these tissular and the free collagen antibodies, irrespective of arthritis susceptibility. These results are against an antibody response restricted to hidden non‐pathogenic epitopes in the resistant mice.
Western Blot Analysis of Human IgG Reactive with the Collagenous Portion of C1q: Evidence of Distinct Binding Specificities
Authors: MÅRTENSSON U. , SJÖHOLM A. G. , STURFELT G. , TRUEDSSON L. , LAURELL A.‐B.
Published: June 2006

An enzyme‐linked immunosorbent assay (ELISA) with purified collagenous C1q fragments in the solid phase was used for detection of C1q‐specific immunoglobulins in the sera of twelve patients with systemic lupus erythematosus (SLE) or the SLE‐like disease hypocomplementemic urticarial vasculitis syndrome (HUVS). By clinical criteria, four patients had SLE, and three HUVS. Five patients had overlap syndromes. All patients demonstrated high concentrations of C1q‐specific IgG and markedly low concentrations of circulating C1q. Detection of C1q‐specific IgG in SLE sera was facilitated by employment of saturating concentrations of collagenous C1q fragments in the solid‐phase ELISA. When added to SLE serum, immune complex‐fixed C1q inhibited binding of IgG to the C1q fragments, whereas addition of C1q alone had limited inhibitory effects. Under similar conditions, using approximately equimolar amounts of C1q relative lo solid‐phase C1q fragments, no ELISA inhibition was obtained after addition of C1q or immune complex‐fixed C1q to a HUVS serum. Even in large excess, purified C1q did not inhibit binding of HUVS‐IgG to solid‐phase C1q fragments. Thus, possible interactions between HUVS‐IgG and native C1q are probably of low affinity. By Western blot analysis. IgG reactive wish the B and C chains of C1q was found in the eight patients with evidence of HUVS, five of whom also showed IgG binding to C′‐C′ and A′‐B′ dimers of collagenous C1q fragments. Sera from SLE patients were negative by Western blot analysis. It seems likely that C1q‐specific IgG in SLE primarily recognizes assembled C1q molecules or collagenous C1q fragments expressing conformational epitopes of bound C1q. Interestingly, patients with evidence of HUVS fairly consistently had zymogen (C1r‐C1s)2 complexes in their serum, while patients with SLE showed high concentrations of complexes containing C1 inhibitor, C1r and C1s. Different binding specificites of C1q‐reactive IgG could be of importance with regard to pathogenetic mechanisms in SLE and HUVS. There was no correlation between findings of C1q‐specific IgG and a variety of autoantibodies associated with SLE and SLF‐like disease.
Human Germinal Centre B Cells Inhibit Mitogen‐Induced proliferation of Mantle Zone B Cells
Authors: RODRÍGUEZ C. , BELLAS C. , BRIEVA J. A.
Published: June 2006

Lymphoid follicles are the main B‐cell areas in peripheral lymphoid tissues. These structures commonly consist of germinal centre (GC) and mantle zone (MZ) regions. In the present work, human tonsillar B cells belonging to these two compartments were purified by a combination of density centrifugation and separation techniques involving the recognition by monoclonal antibodies of specific surface molecules followed by panning and magnetic bead selection. These purified populations were identified as GC and MZ cells by three different criteria: (1) GC cells showed the phenotype IgD‐ CD20+bright CD38+ CD44‐ and peanut agglutinin (PNA)+, and MZ cells were IgD+ CD20+dim CD38‐ CD44+ and PNA‐; (2) morphologically, MZ cells appeared as small resting lymphocytes whereas GC cells consisted of large blastic cells of the germinal centre; (3) functionally, most GC, but not MZ, cells underwent apoptosis early in culture. The isolation of GC and MZ cells allowed the study of their proliferative response. As a result of these studies, GC cells were demonstrated to inhibit the proliferation of MZ cells in response to B‐cell mitogens (Staphylococcus aureus Cowan I and anti‐μ plus BCGF) in a concentration‐dependent way; 50% inhibition was reached at a GC/MZ cell ratio of 1/2. This effect did not require GC‐cell DNA synthesis since similar results were obtained with irradiated GC cells. Neither was it due to a non‐specific toxic effect since GC cells did not alter the proliferative response of autologous T cells to mitogens (phytohaemagglutin and anti‐CD3). The inhibition required cellular contact between GC and MZ cells, and was not restricted by histocompatibility barriers. These data suggest the possible existence of a new regulatory pathway within peripheral B‐cell areas.
C3bi/CR3 is a Main Ligand‐Receptor Interaction in Attachment and Phagocytosis of C3‐Coated Particles by Mouse Peritoneal Macrophages
Authors: KIMURA M. , GRIFFIN F. M.
Published: June 2006

We investigated the relative role of C3bi‐CR3 interaetion in the binding and phagocytosis of EAC43 by mouse peritoneal macrophages. Anti‐Mac‐1 F(ab′)2 markedly inhibited the binding and lymphokine‐induced phagocytosis of both EAC43b and EAC43bi, Fifty per cent inhibition of attachment and phagocytosis occurred at 1 μml of anti‐Mac‐1 F(ab′)2 in the incubation media. On the other hand, ElgG binding and phagocytosis were not inhibited at all even at a concentration of 10 μ/ml. Depletion of divalent ealions from the incubalion media abolished EAC43b and EAC43bi rosettes but not ElgG rosettes or phagocytosis. These data suggested that both EAC43b and EAC43bi binding to macrophages were mediated via CR3.

Because a drastic decrease of FAC43bi rosettes was observed in the case of EAC43bi cells prepared with smaller amounts of C3, a small eontamination of C3bi molecules on EAC43b, itself, cannot explain the efficient attachment of EAC43b, We propose that EAC43b on the macrophage surface can be quickly converted to EAC43bi, forming EAC43bi rosettes, and that those erythrocytes are vigorously ingested by lymphokine‐activated macrophages. In accordance with this hypothesis, we demonstrated that EAC43b was converted to E AC43bi in the medium in whicb macrophages had been ineubated
Effect of Mucosal and Systemic Immunization with Pneumococcal Polysaccharide Type 3, 4 and 14 in the Rat
Authors: VAN DEN DOBBELSTEEN G. P. J. M. , BRUNEKREEF K. , SMINIA T. , VAN REES E. P.
Published: June 2006

Four immunization routes were investigated to induce an immune response against three structurally different types of pneumoccoccal polysaccharide (PPS) in the rat. In particular, the contribution of the IgA isotype in these immune responses was studied. Six days after administration of PPS type 3, 4 or 14. the localization of specific antibody‐containing cells (ACC) in different lymphoid tissues and the antibody titres in serum were studied.

All four routes induced anti‐PPS ACC in the spleen. After intraduodenal, intravenous and especially intraperitoneal administration of PPS, many IgA‐specific anti‐PPS ACC were also found in parathymic and mesenteric lymph nodes and in the lamina propria of intestinal tissue. Several anti‐PPS ACC were found in Peyer's patches, located peripheral of the B‐cell areas. The intratracheal immunization elicited only a local immune response, in bronchus‐associated lymphoid tissues and paratracheal lymph nodes. The localization of these anti‐PPS ACC was influenced by the route of immunization.

After all four investigated routes, specific antibodies were found in serum against PPS. However, some remarkable differences between PPS‐3, 4 and 14 were found in the magnitude of the immune response and the distribution of the isotypes. Both route of immunization and structure of the PPS have a profound influence on the immune responses in rats.
Identification of Immunodominant Antigens during Infection with Mycobacterium tuberculosis
Authors: ANDERSEN P. , ASKGAARD D. , GOTTSCHAU A. , BENNEDSEN J. , NAGAI S. , HERON I.
Published: July 2006

T lymphocytes isolated from mice infected with Mycobacterium tuberculosis respond vigorously to proteins secreted by the bacilli and these antigens may be of importance in the generation of protective immunity against the disease. In this study, short‐term culture filtrate (ST‐CF), which constitutes a complex mixture of secreted proteins, was fractionated by a modified preparative SDS‐PAGE technique. The ability of each fraction to be recognized by T cells isolated from infected mice was evaluated by quantifying proliferation and IFN‐γ production in cell cultures. Two molecular mass regions 4–11 and 26–35 kDa were found to possess marked stimulatory properties. Four potent single antigens were mapped within the stimulatory regions. These purified antigens stimulated T cells isolated from mice at the height of a tuberculous infection to produce large amounts of IFN‐γ, Two of these stimulatory antigens belonged to lhe antigen 85 complex.
Cytokine Production and Responsiveness of Fetal T‐Cell Receptor Vγ3 Thymocytes
Authors: LECLERCQ G. , DE SMEDT M. , PLUM J.
Published: July 2006

The aim of this study was to examine the cytokine production and cytokine responsiveness of the first T‐cell receptor (TcR) positive cells that appear in the murine fetal thymus, namely TcR Vγ3 cells. It is shown that IL‐2‐cultured fetal TcR Vγ3 thymocytes were capable of producing IL‐3, GM‐CSF, TNF‐α and IFN‐γ upon TcR triggering. IL‐2, IL‐4, IL‐5 and IL‐6 could not be detected. With regard to cytokine responsiveness, TcR Vγ3 cells proliferated to a high extent when high concentrations of rIL‐2 were added. rIL‐4or rIL‐7 alone, but not rIL‐1 alone, were capable of inducing a modest proliferation of TcR Vγ3 thymocytes. When combined with low concentrations of IL‐2, a synergistic effect could be observed with IL‐1, IL‐4 or IL‐7. It is shown that the synergistic effect of IL‐2 with IL‐4 was mainly due to induction of IL‐2 receptor expression. The synergistic effect of IL‐2 and IL‐7 on the proliferation of TcR Vγ3 cells could only be partially inhibited by anti‐IL‐2 receptor MoAb, and this antibody had no effcet on the IL‐2 + IL‐1 cultures. These observations can explain the extensive proliferation of TcR Vγ3 thymocytes during fetal life and they indicate that TcR Vγ3 thymocytes have the potential to play a functional role during fetal thymus development.
Oligoclonal T Cells in Rheumatoid Arthritis: Identification Strategy and Molecular Characterization of a Clonal T‐Cell Receptor
Authors: KORTHÄUER U. , HENNERKES B. , MENNINGER H. , MAGES H. W. , ZACHER J. , POTOCNIK A. J. , EMMRICH F. , KROCZEK R. A.
Published: July 2006

Immunodominant antigens in rheumatoid arthritis (RA) should induce an expansion of T cells bearing a corresponding T‐cell receptor (TCR). We therefore analysed the TCR repertoire at the site of inflammation using two fundamentally different strategies. The total TCR repertoire was examined by generating‘representative’T‐cell clone panels, which were subsequently tested for clonality by restriction mapping of the TCRβ gene locus. No clonality was detected in large T‐cell clone panels generated with cells from three patients. However, when we selectively analysed the TCR repertoire of in vivo pre‐activated, interleukin‐2 (IL‐2)‐responsive T cells, significant T‐cell/TCR clonality was found in 2 out or4 patients. The clonal T cells represented a minority of the total T‐cell population with an estimated frequency of 1 in 300 to 1 in 1000 cells. Molecular characterization of a clonal TCR and the use of a specific TCR Vβ MoAb ruled out an over representation of T cells bearing the same Vβlelement in the total T‐cell population, rendering the involvement of superantigens in the induction of T‐cell clonality in this case unlikely.