bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of ultraviolet B irradiation on cell–cell interaction; implication of morphological changes and actin filaments in irradiated cells
Authors: Maekawa T L , Takahashi T A , Fujihara M , Akasaka J , Fujikawa S , Minami M , Sekiguchi S
Published: November 2003

We studied the effects of ultraviolet B (UV-B) irradiation on cell–cell interactions using mouse lymphoma RMA cells and T cell hybridoma HTB-176.10 cells. RMA cells act as stimulators by presenting H-2Kb surface antigens to HTB-176.10 cells, inducing IL-2 production in HTB-176.10 cells. Irradiating RMA cells with 1000 J/m2 UV-B suppressed cell cluster formation between RMA and HTB-176.10 cells and reduced the level of IL-2 production in HTB-176.10 cells, although H-2Kb surface antigens of RMA cells were still expressed. Electron microscopic observations of irradiated RMA cells revealed that UV-B irradiation damaged cell structures, resulting in the disappearance of microvilli on the cell surface, destruction of mitochondria, vacuolation of cytoplasm and swelling of the perinuclear cisterna space. We found that these alterations were accompanied by polymerization of filamentous actin quantified by flow cytometry after NBD-phallacidin staining. Our results suggest that a target of UV-B-induced alterations is actin filaments, which support the cell morphology as the cytoskeleton, and that modification of filamentous actin inhibits interaction between RMA and HTB-176.10 cells. This underlying mechanism may account for the impaired interaction between antigen-presenting cells and T cells after transfusion with UV-B-irradiated allogeneic blood components.
Diverse antigen specificity of erythrocyte-reactive monoclonal autoantibodies from NZB mice
Authors: Desá Oliveira G G , Izui S , Ravirajan C T , Mageed R A K , Lydyard P M , Elson C J , Barker R N
Published: November 2003

The specificities of a panel of erythrocyte-reactive MoAbs derived from NZB mice with autoimmune haemolytic anaemia (AIHA) were determined by immunoprecipitation and immunoblotting. Of the eight antibodies, two (IgG1 MoAb 105-2H and IgG2a MoAb 34-3C) immunoprecipitated a 105-kD component identified as the erythrocyte anion channel band 3. A similar band was also immunoprecipitated by the IgG2b MoAb 34-2B when used at relatively high concentrations, but none of the remaining hybridoma antibodies precipitated any labelled erythrocyte components. In immunoblotting experiments only 34-2B reacted with band 3, indicating that the epitope recognized by this MoAb is robust and differs from the determinant(s) recognized by 105-2H and 34-3C. The remaining MoAbs to react by immunoblotting were the IgM antibodies IE10 and 4C8, both of which bound to a doublet corresponding to band 4.1 from the internal erythrocyte membrane skeleton. Of the three MoAbs which gave negative results in immunoprecipitation and immunoblotting, the IgM antibodies 103-7E and 106-10E reacted poorly with intact erythrocytes by flow cytometry, but the IgG1 antibody 31-9D bound well. ELISAs demonstrated that all four IgM MoAbs are polyreactive, since they bound to histones from a panel of nuclear antigens, and additionally 103-7E reacted with phosphatidyl choline. It is concluded that band 3 is an important autoantigen in NZB AIHA. However, since 3/5 haemolytic MoAbs failed to precipitate this antigen, either these antibodies represent minor components of the total autoantibody response, or responses to diverse possibly non-protein surface antigens also contribute to the pathogenesis of the disease.
IL-12 stimulation but not B7 expression increases melanoma killing by patient cytotoxic T lymphocytes (CTL)
Authors: De Wit D , Flemming C L , Harris J D , Palmer K J , Moore J S , Gore M E , Collins M K L
Published: November 2003

Recent studies have demonstrated that rodent tumour cells engineered to secrete a variety of cytokines, or to express foreign antigens, MHC molecules or co-stimulatory molecules, are rejected by syngeneic animals. These observations have led to the initiation of a number of clinical trials using genetically modified tumour cells, to attempt to stimulate a patient anti-tumour immune response. In this study, a protocol has been developed to test in vitro the specific cytotoxic anti-tumour response generated from melanoma patient lymphocytes. The results showed that IL-12 in combination with IL-2 enhanced the autologous anti-melanoma CTL response, whereas B7.1 antigen expression on tumour cells did not increase anti-melanoma CTL generation. This method could be used to design more appropriate genetically modified tumour vaccines.
Enhanced natural but diminished antibody-mediated cytotoxicity in the lungs of MRLlpr/lpr mice
Authors: Nilsson N , Carlsten H
Published: November 2003

Human systemic lupus erythematosus (SLE) patients, as well as MRLlpr/lpr mice which develop a SLE-like disease, have decreased numbers and functional activity of systemic natural killer (NK) cells. In contrast, it has been found that among lymphocytes recovered from the bronchoalveolar lavage fluid of SLE patients, NK cells were increased in number, correlating with the severity of the lung engagement. The present study was undertaken to assay the capacity for natural killing in the lung compartment of MRLlpr/lpr mice compared with healthy congenic MRL +/+ and heterozygous MRL +/lpr mice. 51Cr-labelled YAC-1 cells were injected intravenously to settle in the lungs where they were targeted for lysis by NK cells. YAC-1 cell killing inversely correlated with radioactivity remaining in the lungs after the assay, and was inhibited by antibody to the asialo-GM1 antigen expressed on NK cells. To analyse the capacity in the lung for cytolysis of non-NK cell-sensitive target cells, a similar in vivo51Cr-release assay was set up for antibody-mediated allospecific cytotoxicity. We demonstrate that MRLlpr/lpr mice throughout their lifespan display significantly increased natural cytotoxic activity in the lungs compared with MRL +/+ and MRL +/lpr mice, as demonstrated by more efficient killing of YAC-1 cells. In contrast, antibody-mediated allospecific cytotoxicity in the lungs was significantly less effective in the MRLlpr/lpr strain.
Inflammatory and immunological markers in preterm infants: correlation with disease
Authors: Jurges E S , Henderson D C
Published: November 2003

Newborn infants often suffer from bacterial and viral infections without presenting typical symptoms. Therefore, reliable methods for detecting and monitoring sepsis in the newborn would be beneficial. In older patients C-reactive protein (CRP) and neopterin have proved useful serum markers of infection and inflammation. Both of these markers are regulated by cytokines, and it has been proposed that cytokines themselves could be used to monitor immune activation and infection. This study has examined the levels of CRP, neopterin, soluble IL-2R, tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) in cord blood samples from both pre-mature and term neonates. Having established reference ranges for these analytes, serial measurements were made in babies requiring intensive care support. The results suggest that in preterm infants the simultaneous measurement of CRP and neopterin, and possibly soluble IL-2R, may provide an accurate early diagnosis of sepsis and may be of use in differentiating between bacterial and viral etiologies. In addition, serial measurement of these markers may help in the early diagnosis of necrotizing enterocolitis (NEC).
Susceptibility to clinically manifest cyclosporine A (CsA)-induced autoimmune disease is associated with interferon-gamma (IFN-γ)-producing CD45RC+RT6− T helper cells
Authors: Beijleveld L J J , Groen H , Broeren C P M , Klatter F A , Kampinga J , Damoiseaux J G M C , Van Breda Vriesman P J C
Published: November 2003

Lethally irradiated Lewis (LEW) rats reconstituted with syngeneic bone marrow and given CsA for a 4-week period, develop, upon withdrawal of CsA, a graft-versus-host-like disease, so-called CsA-induced autoimmunity (CsA-AI). This T cell-mediated autoimmune disease is thymus-dependent; it is generally held that this disease is a consequence of aberrant T cell recovery brought about by CsA. In this study we determined mononuclear cell subsets phenotypically by tri-colour flow cytometry. A strong decrease in recent thymic emigrants (Thy1.1+, TCR αβ+) was observed as a consequence of CsA treatment, eventually resulting in decreased absolute peripheral T cell numbers. In these rats no altered CD4:CD8 T cell ratio was observed before onset of CsA-AI; CD4+ and CD8+ cells consisted predominantly of monocytes (CD4dim+, TCR αβ−) and natural killer cells (CD8+, TCR αβ−), respectively. LEW rats, x-irradiated, syngeneic bone marrow-reconstituted and treated with CsA, showed a marked and persistent, relative expansion of mature CD45RC+, RT6− Th cells. In contrast, Brown-Norway rats treated in a similar fashion, or LEW rats subjected to either CsA treatment or x-irradiation, did not show a comparable expansion of mature CD45RC+, RT6− Th cells, nor did these animals develop CsA-AI. The CD45RC+, RT6− Th cells produced IL-2, and moreover constituted the only Th subset producing IFN-γ upon stimulation, and therefore were considered as Th1-like effector cells. These results are consistent with the view that a persistent preponderance of Th1 cells and not the mere presence of autoreactive cells determines whether or not clinically manifest CsA-AI will occur.
Chronic smoke exposure alters the phenotype pattern and the metabolic response in human alveolar macrophages
Authors: Sköld C M , Lundahl J , Halldén G , Hallgren M , Eklund A
Published: November 2003

Smoking induces a chronic inflammatory process in the lower respiratory tract, where the alveolar macrophages (AM) are the main phagocytes. In the present study, the expressions of different membrane glycoproteins (CD11abc, CD71, CD54, CD14 and CD16) were determined by flow cytometry in AM from smokers and non-smokers after quenching of the intracellular autofluorescence. The metabolic activity of the AM was quantified as a functional test. The expressions of CD11a, CD54 and CD71 were higher in non-smokers' AM than in smokers'. The expressions of CD11b and CD16 were similar between the groups, while the CD11c was higher in smokers' AM compared with non-smokers’. The expression of CD14 was weak in both groups, therefore there was no clear-cut difference between the background and positively labelled cell populations. The metabolic response after in vitro stimulation with the phorbol ester phorbol myristate acetate (PMA) was higher in non-smokers' than in smokers' AM. Our results indicate that chronic exposure to tobacco smoke influences both the expression of AM membrane antigens and the metabolic activity. AM from non-smokers express a phenotype more related to cell proliferation and an accessory function. In contrast, receptors reflecting adhesion and phagocytosis were unaltered or even increased in smokers' AM. The findings suggest a functional change in the AM population after chronic smoke exposure.
Elevation of serum soluble tumour necrosis factor (TNF) receptor and IL-1 receptor antagonist levels in bronchial asthma
Authors: Yoshida S , Hashimoto S , Nakayama T , Kobayashi T , Koizumi A , Horie T
Published: November 2003

The specific inhibitor for TNF-α activity, soluble form of the 55-kD TNF receptor (sTNF-RI) and soluble form of the 75-kD receptor (sTNF-RII), and the specific inhibitor for IL-1 activity, IL-1 receptor antagonist (IL-1Ra), have been identified. It has been shown that the levels of these inhibitors are elevated in plasma/serum and biological fluids in several diseases, and the protective and inhibitory effect of these inhibitors exist in several inflammatory diseases. In the present study, we measured serum levels of sTNF-RI, STNF-RII and IL-1Ra by ELISA in 36 patients with bronchial asthma (16 atopic and 20 non-atopic) during asthma attacks and in stable conditions in order to assess the state of these inhibitors in allergic inflammation. The levels of sTNF-RI, sTNF-RII and IL-1Ra in sera obtained during bronchial asthma attacks were higher than those in sera obtained in stable conditions. These findings were obtained regardless of atopic status. These results suggest that higher levels of serum sTNF-RI, sTNF-RII and IL-1Ra may reflect up-regulation of TNF-R expression and IL-1Ra production in allergic inflammation, and sTNF-RI, sTNF-RII and IL-1Ra may contribute to regulating TNF-α- and IL-1-mediated production and development of allergic inflammation.
Induction of IL-4 by platelet-activating factor
Authors: Huang Y H , Schäfer-Elinder L , Owman H , Lorentzen J C , Rönnelid J , Frostegård J
Published: November 2003

Platelet-activating factor (PAF) is a phospholipid inflammatory mediator which is synthesized by a variety of cells, including monocytes, endothelial cells, mast cells and neutrophils. PAF acts via a recently cloned PAF receptor, present on monocytes and endothelial cells, but not on non-activated lymphocytes. IL-4 is mainly produced by T lymphocytes, and belongs to the Th2 subset of T helper cells. IL-6 is mainly a monocyte/macrophage-derived cytokine with multiple proinflammatory effects. We here report that PAF induces IL-4 production, as determined by ELISPOT. Antibodies to MHC class II inhibited the IL-4 stimulatory effects of PAF. PAF also had the capacity to induce IgA production, as determined by ELISPOT, and IL-6 production in peripheral blood mononuclear cells (PBMC) as determined by ELISA. These PAF-mediated effects were completely inhibited by a specific PAF-receptor antagonist, WEB 2170. Taken together, our data indicate that PAF activates T lymphocytes to IL-4 production by an indirect, monocyte-dependent mechanism dependent on MHC class II. PAF also enhances antibody formation and IL-6 production from PBMC. These findings indicate that PAF activates immune-competent cells, which may be of importance in inflammatory diseases such as asthma, vasculitis and atherosclerosis.
Study of the B cell response to cytochrome P450IID6 in sera from chronic hepatitis C patients
Authors: Parez N , Herzog D , Jacqz-Aigrain E , Homberg J-C , Alvarez F
Published: November 2003

Some patients with chronic hepatitis C develop liver/kidney microsome type 1 antibodies. Some of these autoantibodies are directed against the cytochrome P450IID6. The aim of this study was to characterize the immunogenic sites on the P450IID6 molecule recognized by autoantibodies from individuals infected with the hepatitis C virus. Serum from 24 patients with markers of hepatitis C infection and liver/kidney microsome type 1 antibodies were tested by immunoblotting with human liver microsomal proteins. Eight of them with anti-P450IID6 antibodies were tested with synthetic peptides representing P450IID6 putative immunogenic sites and for their capacity to inhibit in vitro P450IID6 enzymatic activity. Anti-P450IID6 antibodies in the sera of chronic hepatitis C patients were of IgG subclass 1 and in one patient also of IgM type. These sera recognized different P450IID6 peptide sequences consisting of amino acids (aa) 200–214 and aa 271–339. The synthetic peptide between aa 321 and 339 appears to represent the main antigenic site. Five out of eight anti-P450IID6 positive sera were also able to inhibit P450IID6 enzymatic activity in vitro at a dilution of 1 : 100. The anti-P450IID6 autoimmune response in chronic hepatitis C patients is polyclonal, probably inducible, and maintained by the liberation of P450IID6 by hepatocyte lysis. The characterization of the P450IID6 immunogenic site recognized by patients with hepatitis C infection may enable a specific test to be designed to identify those patients with autoimmune hepatitis type 2.
Interaction of human lung surfactant proteins A and D with mite (Dermatophagoides pteronyssinus) allergens
Authors: Wang J Y , Kishore U , Lim B L , Strong P , Reid K B M
Published: November 2003

Human lung surfactant proteins A (SP-A) and D (SP-D) are both collagenous C-type lectins which appear to mediate antimicrobial activity by binding to carbohydrates on micro-organisms and to receptors on phagocytic cells. Purified native SP-A and SP-D, isolated from human bronchoalveolar lavage fluid, were found to bind to whole mite extracts (Dermatophagoides pteronyssinus) and the purified allergen Der p I, in a carbohydrate-specific and calcium-dependent manner. Binding was inhibited by ethylenediamine tetra-acetic acid (EDTA) as well as by maltose in the case of SP-D, or mannose in the case of SP-A. A recombinant polypeptide, which trimerized to form the neck region and carbohydrate recognition domains of SP-D, also inhibited the binding of native SP-D to the whole mite extract and Der p I. Both SP-A and SP-D did not bind to deglycosylated whole mite extracts or to recombinant Der p proteins, which lacked carbohydrate residues. These results suggest that the ability of surfactant proteins to bind certain allergens is mediated through their carbohydrate-recognition domains (CRDs) interacting with carbohydrate residues on the allergens. Moreover, SP-A and SP-D were found to inhibit allergen-specific IgE binding to the mite extracts either via steric hindrance or competitive binding. It is therefore possible that SP-A and SP-D may be involved in the modulation of allergen sensitization and/or the development of allergic reactions.
Increase of tuberculous infection in the organs of B cell-deficient mice
Authors: Vordermeier H M , Venkataprasad N , Harris D P , Ivanyi J
Published: November 2003

Protective immunity against infection with Mycobacterium tuberculosis is imparted by T cells rather than antibodies, but B cells can play a role as antigen-presenting cells and in granuloma formation. We re-evaluated the role of B cells in the course of tuberculous infection in μ-chain knock-out (Ig−) mice. Surprisingly, the organs of M. tuberculosis-infected Ig− mice were found to have three-to eight-fold elevated counts of viable bacilli compared with normal littermates at 3–6 weeks post-infection. Splenic interferon-gamma responses to whole antigen were unimpaired, whilst proliferation to certain mycobacterial peptides was found to be diminished. However, bacille Calmette–Guérin (BCG) vaccination significantly reduced the infection in Ig− mice. The mechanisms by which B cells can influence primary tuberculous infection need further study.
Multiple antigens are altered on T and B lymphocytes from peripheral blood and spleen of patients with Wiskott-Aldrich syndrome
Authors: Gerwin N , Friedrich C , Perez-Atayde A , Rosen F S , Gutierrez-Ramos J C
Published: November 2003

The gene for Wiskott–Aldrich syndrome (WAS) has been recently identified and cloned, but our knowledge of downstream events affected in WAS is limited to a few leucocyte cell surface molecules. To identify cell surface molecules whose abnormal expression could contribute to the functional impairment observed in WAS B and T lymphocytes, we studied the expression of a large panel of antigens on peripheral blood lymphoid cells (PBLC) and on isolated lymphocyte subpopulations from the spleen of WAS patients. WAS T lymphocytes from peripheral blood express increased levels of the activation antigens 4F2, CD49d, CD49e, CD53 and the activation/memory marker CD45RO. In the spleen, however, WAS patients have more CD45RA CD4+ and CD8+ T lymphocytes than normal individuals, suggesting the selective accumulation of presumably naive cells in the WAS spleen. Interestingly, the naive phenotype of the lymphocytes that seem to accumulate in the WAS spleen is confirmed by the absence of increased expression of several activation antigens on their surface, and this correlated with their increased expression of CD43. These lymphocyte abnormalities were accompanied by an abnormal distribution of lymphocyte subsets within the spleen architecture, in particular by the lack of well developed germinal centres and T cell areas. We also found abnormal expression of CD43 and other sialyated proteins such as CDw75 and CD76, whose expression requires the action of specific sialyltransferases. This study shows that the combined impairment in cellular and humoral immunity observed in WAS is the result of multiple molecular abnormalities on the surface of WAS lymphocytes, that in turn might result in recirculation/migration anomalies.
The functional CD40 antigen of fibroblasts may contribute to the proliferation of rheumatoid synovium
Authors: Rissoan M C , Van Kooten C , Chomarat P , Galibert L , Durand I , Thivolet-Bejui F , Miossec P , Banchereau J
Published: November 2003

This paper demonstrates that CD40 is expressed on rheumatoid synovial pannus and primary fibroblast cell lines established from rheumatoid and osteoarthritic synovium as well as normal skin. Among various tested cytokines, interferon-gamma (IFN-γ) and to a lower extent, tumour necrosis factor-alpha (TNF-α) were found to upregulate CD40 expression on fibroblasts. Synovial and skin fibroblasts cultured over CD40 Ligand transfected L cells (L-CD40 L) demonstrate a CD40 specific increase of DNA synthesis as measured by tritiated thymidine incorporation. Cell-cycle analysis and enumeration of viable cells further show that CD40 induced fibroblast proliferation. Costimulation with L-CD40 L and IFN-γ resulted in maximal proliferation. Engagement of fibroblasts CD40 increased the IL-1-induced production of granulocyte macrophage-colony stimulating factor and macrophage inflammatory protein-1α MIP-1α. CD40 L activated fibroblasts showed decreased levels of CD40, but only marginal alterations of other cell-surface antigens. Taken together, the present results indicate that fibroblasts express functional CD40 and suggest a possible role of CD40 L expressing cells, such as activated T cells and mast cells, in the development of synovium hyperplasia.
Efficacy of recombinant granulocyte colony-stimulating factor (rhG-CSF) in experimental colitis
Authors: Hommes D W , Meenan J , Dijkhuizen S , Ten Kate F J W , Tytgat G N J , Van Deventer S J H
Published: November 2003

Inflammatory bowel disease is associated with mucosal neutrophil recruitment and activatation, mediated in part by arachidonic acid metabolites. G-CSF attenuates the immune response to sepsis and ameliorates glycogen storage disease Ib-related colitis. These actions may be effected through the shedding of neutrophil adhesion molecules, or inhibition of proinflammatory mediator synthesis. Immune complex colitis was used to evaluate the effect of rhG-CSF on colonic mucosal inflammation, neutrophil recruitment and the generation of eicosanoids. Immune complex colitis was induced in White New Zealand rabbits. Animals were pretreated with rhG-CSF either 24 h before induction, or at induction, with dosages of 50 and 200 μg/kg. rhG-CSF caused a time- and dose-dependent neutrophilia in all animals. Pretreatment with rhG-CSF resulted in increased tissue myeloperoxidase levels, despite a histologically similar mucosal polymorphonuclear cell infiltrate between treated and control colitis groups. Leukotriene B4 (LTB4) and thromboxane B2 (TXB2) dialysis fluid levels were lower in treated animals, in particular in the groups receiving two doses (LTB4: both P<0.01; TXB2: both P<0.01. Prostaglandin E2 (PGE2) levels in dialysis fluid of the rhG-CSF-treated animals showed no difference from controls. In this model of experimental colitis, high-dose therapy with G-CSF resulted in a marked decrease of proinflammatory mediators, but mucosal generation of the protective PGE2 was preserved. These results suggest that prolonged high-dose therapy with G-CSF may have anti-inflammatory effects in colitis.
Relevance of lymphoproliferative disorders and of anti-C1 inhibitor autoantibodies in acquired angio-oedema
Authors: Cicardi M , Beretta A , Colombo M , Gioffré D , Cugno M , Agostoni A
Published: November 2003

We looked for autoantibodies to C1 inhibitor (C1-INH) and evaluated the relationship of their presence to the associated lymphoproliferative diseases and to the cleaved form of C1-INH in 13 patients with acquired C1-INH deficiency (acquired angio-oedema (AAE)). At the time of manifestation of angio-oedema symptoms or within a few years the following diseases were diagnosed: liver angioma (n = 1), M-components (n = 7, one of whom also had echinococcal liver cysts), breast cancer (n= 1), chronic lymphocytic leukaemia (CLL; n= 1); three patients had no associated disease. Anti-C1-INH autoantibodies, measured both as immunoglobulin binding to C1-INH immobilized onto microtitre plates (ELISA) and as plasma inhibitory activity of C1-INH function, were found in 12 patients. Binding of C1-INH to paraproteins, transferred to Immobilon after agarose gel electrophoresis, was detectable in five of seven M-components associated with AAE. Immunoblotting analysis of SDS–PAGE-separated plasma demonstrated that C1-INH circulated in the cleaved 96-kD form in the 12 patients with autoantibodies, but not in the one without. In conclusion, the large majority of our patients have autoantibodies to C1-INH. Circulating autoantibodies are necessary for the generation of cleaved C1-INH. The paraproteins associated with AAE are frequently autoantibodies to C1-INH and thus account for its consumption.
Antibodies to membranes of endothelial cells and fibroblasts in scleroderma
Authors: Hill M B , Phipps J L , Cartwright R J , Milford Ward A , Greaves M , Hughes P
Published: November 2003

Anti-endothelial and other cell membrane-reactive antibodies in scleroderma were characterized by immunoblotting sera with membrane and cytosol preparations of human umbilical vein endothelial cells (HUVEC), dermal fibroblasts and a T cell lymphoma HUT78. Antibodies reactive with HUVEC membranes were found in 17 of 20 patients with scleroderma (33 bands) in contrast to only two of 20 controls (two bands; P<0.01) and three of 11 patients with myocardial infarction (four bands). Eleven of the 20 patients possessed antibodies that were specific for HUVEC membrane and did not cross-react with other cell lines. Analysis of patient subgroups showed that HUVEC membrane antibodies were present in nine of 11 patients with systemic sclerosis and in all nine with the CREST syndrome, and were HUVEC-specific in five and six of these cases, respectively. Although considerable heterogeneity was seen, antibodies to an 18–19-kD membrane epitope were found in 11 of the 20 patients but in none of the controls (P<0.01). This antibody which reacted particularly with HUVEC (n=9) and HUT78 membranes (n=9) was associated with CREST syndrome rather than systemic sclerosis (9/9 versus 1/11; P<0.01), and after elution was shown to possess anticentromere activity. In addition, antibodies reactive with both fibroblast (n=11; 18 bands) and HUT78 membranes (n=18; 42 bands) were detected and were specific for either fibroblast or HUT78 membranes in nine and 14 patients, respectively. There was no significant difference in the incidence of these fibroblasts and HUT78 membrane antibodies in the two patient subgroups. These findings support the concept that membrane-reactive antibodies, including anticentromeric antibodies, may play a central role in the pathogenesis of scleroderma, through their ability to react with endothelial cells.
Contribution of tumour necrosis factor-alpha (TNF-α) in host defence mechanism against Cryptococcus neoformans
Authors: Kawakami K , Qifeng X , Tohyama M , Qureshi M H , Saito A
Published: November 2003

We investigated the role of TNF-α in the host defence mechanism against infection with a virulent strain of Cryptococcus neoformans. Administration of exogenous recombinant human TNF-α significantly prolonged the survival time of mice infected by intratracheal instillation of the organism. Surprisingly, neutralizing MoAb to murine TNF-α did not shorten their survival time, a finding inconsistent with previous results. To investigate the cause of this inconsistency, we examined the production of TNF-α in the lungs of infected mice. During the course of cryptococcosis, there was little or no generation of TNF-α mRNA in the lung. This might be partly due to a direct inhibitory action of the fungal microorganism on TNF-α production by macrophages. In vitro production of TNF-α by murine interferon-gamma (IFN-γ)- and lipopolysaccharide (LPS)-stimulated macrophages was strongly inhibited by co-culturing with the whole yeast cells. In contrast, administration of recombinant murine IL-12 markedly induced TNF-α production and the neutralizing anti-TNF-α MoAb strongly blocked IL-12-induced protection of mice against cryptococcal infection. These results indicate that endogenously synthesized TNF-α has the potential to contribute to the elimination of C. neoformans and partly mediates the protective effect of IL-12.
Dynamics of proinflammatory cytokine expression in the joints of mice with collagen-induced arthritis (CIA)
Authors: MARINOVA-MUTAFCHIEVA L , WILLIAMS R O , MASON L J , MAURI C , FELDMANN M , MAINI R N
Published: November 2003

This report contains a description of the cellular localization and kinetics of proinflammatory cytokine expression in murine CIA, a model for rheumatoid arthritis. Tissue cryostat sections of undecalcified paws from type II collagen-immunized DBA/1 mice, taken 1–10 days after the onset of clinical arthritis, were examined for the presence of tumour necrosis factor-alpha (TNF-α), IL-1β and IL-6 using an indirect immunoperoxidase technique. In parallel, interferon-gamma (IFN-γ) production by lymph node cells, stimulated in vitro with type II collagen, was assessed as a marker of T cell activity. The main areas of TNF-α, IL-1β and IL-6 expression were in the synovial lining layer and in tissue contiguous with cartilage and bone (the marginal zone), in particular at sites of pannus formation and joint erosion. There was a progressive increase in the number of TNF-α-, IL-1β- and IL-6-positive cells from day 1 to day 10 of arthritis, during which time IFN-γ production by CD4+ T cells from draining lymph nodes declined sharply. A further finding of potential significance was that TNF-α was consistently detected at day 1 of arthritis, whereas IL-1β-positive cells were not found until day 3, suggesting that the expression of TNF-α precedes that of IL-1β.
Adhesion molecule expression in vivo on extraocular muscles (EOM) in thyroid-associated ophthalmopathy (TAO)
Authors: PAPPA A , CALDER V , FELLS P , LIGHTMAN S
Published: November 2003

TAO is an autoimmune condition characterized by mononuclear cell infiltration of the extraocular muscles (EOM) and/or the orbital fat/connective tissue with associated deposition of glycosaminoglycans (GAG) in the interstitial spaces. In this study, the presence and distribution of the vascular adhesion molecules intercellular adhesion molecule-1 (ICAM-1), endothelial-leucocyte adhesion molecule-1 (ELAM-1), vascular cell adhesion molecule-1 (VCAM-1) and the leucocyte integrins CD11a/CD18, CD11b/CD18, CD11c/CD18 were investigated. Nineteen EOM biopsies were collected from 17 patients with early (n = 6) and late (n = 13) TAO as well as from 12 non-TAO control patients. Consecutive cryostat sections of these biopsies were immunostained with MoAbs to the above-mentioned molecules and haematoxylin and eosin. Primary antibody binding was visualized using an avidin-biotin system. In early untreated TAO specimens, the interstitial and perimysial connective tissue surrounding EOM fibres and numerous mononuclear cells stained strongly for ICAM-1. In contrast, the vascular endothelial cells (ulex lectin-positive) stained strongly for ELAM-1 (E-selectin), VCAM-1 as well as ICAM-1. In late disease, the same distribution of immunoreactivity for ICAM-1, ELAM-1 and VCAM-1 was observed, but with significantly lower staining. The leucocyte integrins (CD11a, CD11b, CD11c) were again expressed at significantly higher levels in early TAO specimens compared with late TAO specimens and were minimal or absent in the EOM biopsies harvested from control patients. In conclusion, increased expression of adhesion molecules studied correlated with early active disease and was reduced in later stages.
Different modulation by histamine of IL-4 and interferon-gamma (IFN-γ) release according to the phenotype of human Th0, Th1 and Th2 clones
Authors: LAGIER B , LEBEL B , BOUSQUET J , PÈNE J
Published: November 2003

Histamine, an important inflammatory mediator in allergic diseases and asthma, has been reported to have modulator effects on T cells, suggesting that the bronchial microenvironment may regulate the function of resident T cells. We examined the effect of histamine on the release of the Th2-associated cytokines IL-4 and IL-5 and the Th1-associated cytokine IFN-γ by 30 CD4+ T cell clones from peripheral blood or bronchial biopsy of one atopic subject. Based on the IL-4/IFN-γ ratio, the clones were ascribed to the Th2 (ratio >1), Th0 (ratio  0.1 and 1) or Th1 (ratio <0.1) phenotype. Histamine inhibited IFN-γ production by Th1-like cells (P<0.02, Kruskall–Wallis), especially from bronchial biopsy, but had no effect on IL-4 release. Regarding Th0 clones, histamine inhibited IL-4 production (P<0.02) in a dose-dependent manner and slightly inhibited IFN-γ production, but had no effect on Th2-like cells. Histamine had a heterogeneous and insignificant effect on IL-5 production. The H2-receptor antagonist ranitidine completely reversed the inhibition of IL-4 and IFN-γ production, whereas the agonist dimaprit mimicked this effect. In contrast, H1- and H3-receptor agonists and antagonists had no significant effect. These data demonstrate that histamine has different effects on IL-4 and IFN-γ release by T helper cells according to their phenotype via H2-receptors. This study extends the immunomodulatory effects of histamine which may contribute to the perpetuation of airway inflammation in asthma.
Normal CD5+ B lymphocytes are poor stimulators in the autologous mixed lymphocyte reaction (AMLR)
Authors: FERNANDEZ L A , CHENG R , SADEK I , NEVILL T J , MACSWEEN J M
Published: November 2003

The AMLR is decreased in chronic lymphocytic leukaemia (CLL), which is characterized by a monoclonal expansion of CD5+ B lymphocytes. Since B cells are used to stimulate the AMLR, we investigated the capacity of normal CD5+ B cells to function as stimulators in the AMLR. We isolated B cells from the peripheral blood of normal individuals and fractionated them into subpopulations enriched for CD5+ and CD5− cells, which were used as stimulators in the AMLR. We found that the CD5− B cells were excellent stimulators, whereas stimulation of AMLR by B cells enriched for CD5+ cells resulted in significantly reduced proliferative responses (P<0.005). This suggests that the reduced AMLR in CLL is due to the predominance of CD5+ B lymphocytes in the stimulating cell population.
Difficulties in the ascertainment of C9 deficiency: lessons to be drawn from a compound heterozygote C9-deficient subject
Authors: HOBART M J , FERNIE B A , WÜRZNER R , OLDROYD R G , HARRISON R A , JOYSEY V , LACHMANN P J
Published: November 2003

A group of patients with long-surviving mismatched kidney allografts were investigated for complement function using haemolytic assays in agarose gels. One patient was found to have no alternative pathway activity but a low normal classical pathway. Surprisingly, investigation revealed that the patient's complement was normal for all components except C9, which was functionally absent. The patient was shown to be heterozygous for DNA markers in the C6, C7 and C9 region of chromosome 5 and therefore appears to be a compound heterozygote for two uncharacterized C9 deficiency genes. Serological analysis by ELISA revealed that he has trace concentrations of a non-functional C9 molecule. Western blot analysis was not sufficiently sensitive to permit detection of this molecule. We hypothesize that the patient is heterozygous for a complete deficiency of C9 and for a gene directing hyposynthesis of a defective C9. We also suggest that C9 deficiency may be more common among Caucasians than has been reported.
Regulatory role of exogenous IL-10 in the development of immune response versus Cryptococcus neoformans
Authors: MONARI C , RETINI C , PALAZZETTI B , BISTONI F , VECCHIRELLI A
Published: November 2003

The most important event involved in host defence against Cryptococcus neoformans is the development of an adequate cell-mediated immune response. IL-10, abundantly produced during AIDS progression, could be a negative factor that affects the T cell response through its own immunosuppressive action on antigen-presenting cells. To determine whether this cytokine affects the course of immune response against C. neoformans, we added exogenous IL-10 to cultured Cryptococcus-laden monocytes plus T lymphocytes. The data from this study confirmed the down-regulatory effect of exogenous IL-10 on monocytes and expanded the known inhibitory role to include an increase of the deleterious effect due to capsular material of C. neoformans on (i) lymphoproliferation, (ii) down-regulation of MHC class II molecules, (iii) inhibition of IL-2 mRNA expression and protein secretion by T lymphocytes. These results indicate that the presence of IL-10 in AIDS patients, due to the progression of disease, could represent a pivotal problem contributing to augment the pathogenic effect of C. neoformans.
Genetic differences in immune reactivity to mercuric chloride (HgCl2): immunosuppression of H-2d mice is mediated by interferon-gamma (IFN-γ)
Authors: DOTH M , FRICKE M , NICOLETTI F , GAROTTA G , VAN VELTHUYSEN M-L , BRUIJN J A , GLEICHMANN E
Published: November 2003

Upon treatment with HgCl2, H–2s mice, such as B10.S, develop an activation of B lymphocytes that depends, at least partially, on activation of T helper type 2 (Th2) cells and results in increased serum levels of IgG1 and IgE, appearance of IgG autoantibodies, and development of immune glomerulonephritis and vasculitis. Results of previous studies and of experiments presented here indicate that the B cell activation and systemic autoimmune disease fail to develop in MHC-congenic B10.D2 (H-2d) and B10.BR (H–2k) mice treated with HgCl2, although B10.D2 T cells showed signs of activation by and specificity for HgCl2 comparable to those seen in strain B10.S. Here, we report that following HgCl2 injections the antibody response to sheep erythrocytes is normal in B10.S, but suppressed in B10.D2 mice. This suppression was prevented by MoAb to mouse IFN-γ. Conversely, treatment of B10.D2 mice with murine recombinant IFN-γ (rIFN-γ) was able to reproduce the immunosuppression seen after HgCl2 treatment. In B10.S mice, it took administration of both rIFN-γ and HgCl2 to suppress the anti-sheep erythrocyte response. Although rIFN-γ diminished the increase in IgE serum levels of HgCl2-treated B10.S mice, it failed to prevent their autoantibody production and immune glomerulonephritis. These findings further strengthen the concept that B10.S mice react to HgCl2 by preferential activation of their Th2 cells producing IL-4, whereas B10.D2 mice react to HgCl2 by preferential activation of their Th1 cells, which produce IFN-γ and thus suppress antibody responses.
Evidence for genetic heterogeneity in inflammatory bowel disease (IBD); HLA genes in the predisposition to suffer from ulcerative colitis (UC) and Crohn's disease (CD)
Authors: BOUMA G , OUDKERK POOL M , CRUSIUS J B A , TH SCHREUDER G M , HELLEMANS H P R , MEIJER B U G A , KOSTENSE P J , GIPHART M J , MEUWISSEN S G M , PEÑA A S
Published: November 2003

Family and epidemiological studies support a genetic susceptibility to UC and CD. Conflicting reports regarding associations between UC and HLA-DR2 and between CD and various HLA alleles have been published. The aim of this study was to determine whether molecularly defined HLA-DR genes are associated with these diseases in a Dutch group of patients. Fifty-nine unrelated Dutch UC patients and 89 CD patients were typed using DNA-based methods. A total of 2400 healthy local blood donors served as controls. The phenotype frequency of the HLA-DRB1*15 allele was increased in UC patients compared with controls (42% versus 26% in controls; P = 0.006; odds ratio (OR) = 2.1), and was predominantly found in female patients (53% versus 24%; P = 0.001; OR = 3.5). The DRB1*15 allele was increased in UC patients having a positive family history (P = 0.01; OR = 5.8). Among the 16 patients who showed an increase in extent of disease during follow up, 10 were DRB1*15+ (P = 0.002; OR = 4.8). The frequency of the DRB1*13 allele was decreased in patients with UC (15% versus 28% in controls; P = 0.04; OR = 0.5). In CD, no association was observed between disease or particular clinical subgroups and any allele tested. The present study provides additional evidence for the genetic association between UC and HLA-DRB1*15, and supports recent findings that the susceptibility gene(s) for CD is not located in the HLA class II region.
Specificity and immunochemical properties of antibodies to bacterial DNA in sera of normal human subjects and patients with systemic lupus erythematosus (SLE)
Authors: WU Z-Q , DRAYTON D , PISETSKY D S
Published: November 2003

To elucidate the mechanisms of anti-DNA production, we assessed the binding of sera of normal human subjects (NHS) and patients with SLE to a panel of bacterial and mammalian DNA. Using single-stranded DNA as antigens in an ELISA, NHS showed significant binding to some but not all bacterial DNA, while lacking reactivity to calf thymus DNA. Among bacterial DNA, the highest levels of binding were observed with DNA from Micrococcus lysodeikticus and Staphylococcus aureus. In contrast, SLE sera showed high levels of binding to all DNA tested. To evaluate further immunochemical properties of the anti-DNA antibodies, the subclass distribution of these responses was evaluated by subclass-specific reagents. While NHS showed a predominance of IgG2 antibodies to bacterial DNA, SLE sera had a predominance of IgG1 antibodies to these antigens. Together, these results provide further evidence for the antigenicity of bacterial DNA and suggest that NHS and SLE anti-DNA differ in the patterns of epitope recognition as well as mechanisms of induction.
Molecular aberrations in the MHC class I-restricted pathway for antigen presentation in methylcholanthrene sarcomas from nude mice: discrepancies between MHC mRNA and surface protein
Authors: ENGEL A-M , SVANE I M , MADSEN M W , PEDERSEN M , WERDELIN O
Published: November 2003

In a previous study, we demonstrated that eight sarcomas induced by chemical carcinogenesis in nude mice were rejected by syngeneic immunocompetent recipients at a much higher rate than eight sarcomas induced with the same method in syngeneic immmunocompetent mice. In the present study, we investigated these 16 sarcomas for structural and quantitative aberrations in components of the MHC class I-restricted antigen-processing and -presentation pathway. Considerable discrepancies between mRNA levels and cell surface protein expression of MHC class I (Kd, Dd and Ld) molecules were observed almost exclusively in the tumours derived from nude mice. Several of the nude mouse-derived tumours also displayed incongruent levels of heavy chain mRNA and β2-microglobulin mRNA. These findings are taken as indications of abnormal regulation of gene transcription in nude mouse tumours, and if this abnormal regulation extends to the entire genome, it may explain the pronounced immunogenicity of these tumours. Proteasome composition, heat shock protein expression, TAP-molecule inducibility and intercellular adhesion molecule-1 expression were investigated in the same tumours. We found no indications of structural defects or quantitative differences in these molecules between the two groups of tumours.
Characterization of proliferative responses and cytokine mRNA profiles induced by Vespula venom in patients with severe reactions to wasp stings
Authors: BONAY M , ECHCHAKIR H , LECOSSIER D , LAINÉ P , HERMAN D , HANCE A J , SOLER P , TAZI A
Published: November 2003

The reasons why severe allergic reactions to bee and wasp stings develop in only a small portion of exposed individuals are incompletely understood, but differences in T cell responses to venom antigens comparing allergic and non-allergic individuals are likely to be important. To identify such differences, venom-induced proliferative responses and cytokine mRNA production by blood mononuclear cells from Vespula venom-allergic patients and non-allergic individuals were compared. Mononuclear cells from most venom-allergic patients proliferated in response to alkylated Vespula venom (7275 ± 8387 ct/min, n = 19), and the extent of proliferation was greater for patients with a history of multiple prior stings and those with high levels of venom-specific IgE. Although mononuclear cells from non-allergic subjects showed little or no proliferation in response to venom (926 ± 711 ct/min, n = 8), production of mRNAs coding for IL-2, IL-4, IL-5, IL-10 and interferon-gamma (IFN-γ) in response to Vespula venom by cells from non-allergic subjects was detected by reverse transcriptase-polymerase chain reaction (RT-PCR), indicating that these individuals had been previously sensitized to venom antigens. In contrast to the Th0 cytokine mRNA profile observed for non-allergic individuals, venom-allergic patients released a more restricted profile of cytokines following stimulation with venom. Only IFN-γ mRNA expression was detected in all individuals evaluated, whereas IL-2 mRNA was not detected during the first 48 h of stimulation, and T cells from only one of three venom-allergic individuals produced detectable IL-4 or IL-5 mRNA. The difference in cytokine profiles observed comparing venom-allergic patients and non-allergic controls could not be attributed to intrinsic differences in T cells from these individuals, because polyclonal stimulation with phorbol myristate acetate (PMA) + ionophore induced similar cytokine mRNA profiles in the two groups. These studies demonstrate clear differences in the T cell responses of venom-allergic subjects, that may contribute to the development of severe allergic reactions in these individuals.
Effector molecules of the host defence mechanism against Mycobacterium avium complex: the evidence showing that reactive oxygen intermediates, reactive nitrogen intermediates, and free fatty acids each alone are not decisive in expression of macrophage antimicrobial activity against the parasites
Authors: TOMIOKA H , SATO K , SANO C , AKAKI T , SHIMIZU T , KAJITANI H , SAITO H
Published: November 2003

In this study, we evaluated the roles of reactive oxygen intermediates (ROI), reactive nitrogen intermediates (RNI), and free fatty acids (FFA) as effectors of the macrophage-mediated host defence mechanism against Mycobacterium avium complex (MAC). First, M. avium (three strains) and M. intracellulare (two strains) were treated with the H2O2-Fe2+-mediated halogenation system, acidified NaNO2-derived RNI, or FFA (linolenic acid) in sodium acetate buffer pH 5.5, and then counted for the number of residual colony-forming units (CFU) of organisms. Although these effectors exerted strong bactericidal activity against the MAC, the susceptibility of test organisms markedly varied from strain to strain. There was no significant relationship between the degree of resistance of a given MAC strain to these effectors and its virulence in mice, indicating that ROI, RNI, and FFA each alone are not decisive as the effector components of the host defence mechanism against the MAC. Second, the increase in ROI-producing ability in murine peritoneal macrophages due to tumour necrosis factor-alpha (TNF-α) treatment was not accompanied by parallel potentiation of anti-MAC activity of the same macrophage population. This excludes the possibility that ROI play a central role in macrophage-mediated killing and inhibition of MAC organisms. Third, anti-MAC activity of BAM3 macrophage cell line was not significantly attenuated by NG-monomethyl-l-arginine (NO synthase-inhibitor causing reduction of RNI production) or by quinacrine (phospholipase A2-inhibitor causing reduction of FFA release), indicating that RNI and FFA each alone do not play crucial roles in the expression of macrophage antimicrobial activity against the MAC. The present findings suggest important roles of collaborating actions of various antimicrobial effectors and/or the participation of other kinds of effectors in macrophage-mediated killing and inhibition of MAC organisms.