bluuebunny/crossref_metadata_2025 huggingface dataset:
IgG subclass reactivity to human cardiac myosin in cardiomyopathy patients is indicative of a Th1-like autoimmune disease
Authors: SKYLLOURIOTIS P , SKYLLOURIOTIS-LAZAROU M , NATTER S , STEINER R , SPITZAUER S , KAPIOTIS S , VALENT P , HIRSCHL A M , GUBER S E , LAUFER G , WOLLENEK G , WOLNER E , WIMMER M , VALENTA R
Published: March 2003

Studies performed in mice together with the demonstration of increased levels of heart-specific autoantibodies, cytokines and cytokine receptors in sera from cardiomyopathy (CMP) patients argued for a pathogenic role of autoimmune mechanisms in CMP. This study was designed to analyse the presence of IgG anti-heart antibodies in sera from patients suffering from hypertrophic and dilatative forms of CMP as well as from patients with ischaemic heart disease and healthy individuals. Patients' sera were analysed for IgG reactivity to Western-blotted extracts prepared from human epithelial and endothelial cells, heart and skeletal muscle specimens as well as from Streptococcus pyogenes. The IgG subclass (IgG1–4) reactivity to purified human cardiac myosin was analysed by ELISA. While sera from CMP patients and healthy individuals displayed comparable IgG reactivity to a variety of human proteins, cardiac myosin represented the prominent antigen detected strongly and preferentially by sera from CMP patients. Pronounced IgG anti-cardiac myosin reactivity was frequently found in sera from patients with dilatative CMP and reduced ventricular function. ELISA analyses revealed a prominent IgG2/IgG3 anti-cardiac myosin reactivity in CMP sera, indicating a preferential Th1-like immune response. Elevated anti-cytomegalovirus, anti-enterovirus IgG titres as well as IgG reactivity to nitrocellulose-blotted S. pyogenes proteins were also frequently observed in the group of CMP patients. If further work can support the hypothesis that autoreactivity to cardiac myosin represents a pathogenic factor in CMP, specific immunomodulation of this Th1- towards a Th2-like immune response may represent a promising therapeutic strategy for CMP.
Inhibitory effects of transforming growth factor-beta (TGF-β) on certain functions of intraepithelial lymphocytes
Authors: EBERT E C
Published: March 2003

Human intraepithelial lymphocytes (IEL), CD8+ lymphocytes located between epithelial cells, are likely to be influenced by the immunosuppressive cytokine, TGF-β, secreted by epithelial cells. This study evaluates the effects of TGF-β on IEL functions. IEL were derived from proximal jejunum of patients undergoing gastric bypass operations for morbid obesity. Proliferation was determined by 3H-thymidine incorporation; IL-2 production, by ELISA; expression of IL-2 receptor, CD2, HML1, CD16, and CD56, by immunofluorescence; binding, by adherence of radiolabelled cells; and cytotoxicity by 51Cr-release assay. TGF-β (≥ 1 ng/ml) inhibited the mitosis of IEL to mitogens, IL-7, and stimuli of the CD2 and CD3 pathways. The blocking effect did not target the activation events of IL-2 production and receptor generation. Rather, it reduced cell division after activation when added 24 h after initiating the culture. Antibody neutralization of naturally occurring TGF-β increased IEL proliferation to IL-2, but not to the other stimuli. Of the multiple surface markers tested, only CD2 and HML1 expression increased with TGF-β and decreased with antibody to TGF-β, although the cytokine and the neutralizing antibody had no effects on IEL binding to colon cancer. TGF-β reduced the number of CD56+ IEL and the lymphokine-activated killing when co-cultured with IL-7 but not with IL-2 or IL-15. TGF-β inhibits certain IEL functions: the reduction in cell division rather than activation and a decline in IL-7-mediated lysis of colon cancer due to a lowering of the number of natural killer cells.
Inhibitory effects of anti-rheumatic drugs on vascular endothelial growth factor in cultured rheumatoid synovial cells
Authors: NAGASHIMA M , YOSHINO S , AONO H , TAKAI M , SASANO M
Published: March 2003

Vascular endothelial growth factor (VEGF) is a potent inducer of angiogenesis and is constitutively expressed in the synovium of rheumatoid arthritis (RA). Over-expression of VEGF may play an important role in pathogenic vascularization and synovial hyperplasia of RA. In the present study, we examined whether disease-modifying anti-rheumatic drugs (DMARDs), including bucillamine (BUC), gold sodium thiomalate (GST), methotrexate (MTX) and salazosulfapiridine (SASP), act by inhibiting the production of VEGF by cultured synovial cells of patients with RA. Treatment of cultured synoviocytes with lipopolysaccharide (LPS) significantly increased VEGF production by cultured synovial cells. BUC significantly inhibited LPS-induced VEGF production, while GST tended to inhibit the production of VEGF. The inhibitory effects on VEGF production were dose-dependent. In contrast, MTX and SASP did not affect VEGF production. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis showed that BUC also inhibited LPS-induced VEGF mRNA expression in RA synovial cells. The present study provides the first evidence that BUC inhibits VEGF production and the expression of its mRNA in synovial cells of RA patients. Our results indicate that the anti-rheumatic effects of BUC are mediated by suppression of angiogenesis and synovial proliferation in the RA synovium through the inhibition of VEGF production by synovial cells.
Leishmania-specific T cells expressing interferon-gamma (IFN-γ) and IL-10 upon activation are expanded in individuals cured of visceral leishmaniasis
Authors: KEMP K , KEMP M , KHARAZMI A , ISMAIL A , KURTZHALS J A L , HVIID L , THEANDER T G
Published: March 2003

Peripheral blood mononuclear cells (PBMC) from patients who have recovered from visceral leishmaniasis often respond to Leishmania antigens in vitro by production of both IL-4, IFN-γ and IL-10. In order to establish the cellular sources of these cytokines, we activated cells from individuals with a history of visceral leishmaniasis with Leishmania antigen for 6 days in culture, and identified cytokine production at the single-cell level by flow cytometry. The cytokines were only found in CD3+ cells and among these mainly within the CD4+ subset. The percentage of cytokine-producing cells was compared in Leishmania-activated PBMC cultures from the previous patients and from individuals living in a village where leishmaniasis does not occur. The percentage of IL-10- and IFN-γ-containing cells was significantly higher in the previous patients than in the controls, indicating that Leishmania-specific T cells producing IL-10 and/or IFN-γ had been expanded as a result of the infection. The cytokine-producing cells in the previous patients could be divided into three types: (i) cells producing IFN-γ only; (ii) cells producing IL-4 only; and (iii) cells producing IFN-γ and IL-10 simultaneously. The first and second group of cells can be described as Th1- and Th2-type cells, respectively. The third group could be a regulatory subset of T cells important for maintaining a balance between Th1- and Th2-type cells in these individuals.
Regulation of the expression of Fcγ receptor on circulating neutrophils and monocytes in Kawasaki disease
Authors: Nakatani K , Takeshita S , Tsujimoto H , Kawamura Y , Kawase H , Sekine I
Published: March 2003

To investigate the regulation of Fcγ receptor (FcγR) expression on circulating phagocytes in Kawasaki disease (KD), we analysed the expressions of FcγRI, II and III on neutrophils and monocytes in 20 patients with KD, 10 with a bacterial infection (BI), 10 with a viral infection (VI), and 10 healthy controls (HC) using flow cytometric analysis. The KD patients had a significantly higher level of FcγRI expression on neutrophils, but not on monocytes, than the BI, VI and HC patients. FcγRII expression on neutrophils was significantly higher in KD, BI and VI than HC, but there was no significant difference in FcγRII expression among KD, BI and VI. FcγRIII expression on neutrophils in KD was significantly lower than in VI and HC, but was higher on monocytes. A kinetic analysis of FcγR expression in KD demonstrated the expression of FcγRI and II on neutrophils to decline, but no remarkable change was observed in the monocytes, from the subacute phase through the convalescent phase. In addition, FcγRIII expression on neutrophils increased, while FcγRIII expression on monocytes decreased during the time course of KD. FcγR expression in the acute phase of KD is thus characterized by markedly increased expression of FcγRI on neutrophils, followed by a subsequent decrease, and decreased expression of FcγRIII on neutrophils and increased expression of FcγRIII on monocytes followed by a reverse kinetics during the clinical course. These findings are thus considered to reflect the functional up-regulation of neutrophils and monocytes in KD.
The T cell receptor repertoire of CD8+CD28− T lymphocytes is dominated by expanded clones that persist over time
Authors: Mugnaini E N , Egeland T , Spurkland A , Brinchmann J E
Published: March 2003

The costimulatory molecule CD28 is expressed on almost all CD4+ T cells, but on only a portion of CD8+ T cells in healthy human adults. αβ T cells may thus be divided into three phenotypically and functionally different subsets: CD4+, CD8+CD28+ and CD8+CD28−. Using peripheral blood lymphocytes from six healthy adults, we have studied the T cell receptor (TCR) repertoire within these subsets by analysis of the distribution of lengths of the complementarity determining region 3 (CDR3) of the beta variable (BV) transcripts and flow cytometric analysis of TCR Vβ usage. Expanded CDR3 lengths were identified in 86% of BV families within CD8+CD28− T cells, but in only 4% within CD4+ T cells, and 35% within CD8+CD28+ T cells (P < 0.01). When sequenced, the majority of expanded peaks were found to be dominated by single clones. Identical expanded clones were found within both CD8+CD28+ and CD8+CD28− subsets, consistent with the belief that CD8+CD28− T cells descend directly from CD8+CD28+ T cells. Greatly expanded CD28− clones were found within both CD8+ and CD4+ subsets and persisted at the same magnitude for up to 4.5 years of observation. The finding of a small proportion of cells expressing Ki-67 showed that some of these clonally expanded cells were in the active stages of the cell cycle, but few of the cells expressed activation markers CD69, CD25, CD71 or CD122. One likely explanation for the persistence of expanded peripheral lymphocyte populations in healthy individuals is the presence of persistent antigen.
The influence of age and gender on serum dehydroepiandrosterone sulphate (DHEA-S), IL-6, IL-6 soluble receptor (IL-6 sR) and transforming growth factor beta 1 (TGF-β1) levels in normal healthy blood donors
Authors: Young D G , Skibinski G , Mason J I , James K
Published: March 2003

Dysregulation of IL-6 synthesis is thought to play a role in the development of a number of age-related conditions, such as rheumatoid arthritis, osteoporosis, atherosclerosis, Alzheimer's disease and B cell malignancies. Recently it has been suggested that the production of IL-6 is influenced by the adrenal hormone dehydroepiandrosterone (DHEA) and its sulphated derivative DHEA-S. In humans we investigated the relationship between DHEA-S, IL-6, IL-6 sR and TGF-β1 in the serum of normal healthy male and female blood donors. Using immunoassay techniques we found that the serum levels of DHEA-S significantly (P = 0.0001) decreased with age in both males and females. Furthermore, mean DHEA-S levels in all age groups were significantly (P = 0.0001) higher in males. Such correlations were not apparent for IL-6 using a standard assay, but a high sensitivity assay revealed that serum IL-6 was significantly (P = 0.0018) positively correlated with age in males only. In addition, serum levels of DHEA-S were significantly (P = 0.048) negatively correlated with serum IL-6, again in male subjects only. In contrast, serum IL-6 sR and TGF-β1 levels were not correlated with age in either males or females and were not significantly different between the sexes. However, a significant (P = 0.024) negative correlation between DHEA-S and IL-6 sR was found in males. These studies clearly highlight the complex nature of the relationship between these molecules in the ageing process in normal healthy blood donors and demonstrate the need to use high sensitivity assays when measuring IL-6 in apparently healthy individuals under the age of 70 years.
Release of CXC-chemokines by human lung microvascular endothelial cells (LMVEC) compared with macrovascular umbilical vein endothelial cells
Authors: Beck G , Yard B A , Breedijk A J , Van Ackern K , Van Der Woude F J
Published: March 2003

In the present study, the sensitivity of LMVEC and human umbilical vein endothelial cells (HUVEC) to lipopolysaccharide (LPS) and the proinflammatory cytokines IL-1, tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) was compared. To this end, the production of the CC- (MCP-1), CXC- (IL-8, ENA-78, Groα, NAP-2, GCP-2) and CX3C (fractalkine) chemokines was studied. A low basal production of these chemokines was observed in both cell types. TNF-α, IL-1 and LPS up-regulated all chemokines tested. IFN-γ however was only able to up-regulate MCP-1 production. LMVEC were more sensitive to IL-1 and LPS compared with HUVEC, since LMVEC produced significantly more MCP-1, ENA-78 and Groα (P < 0.01) under these conditions. Maximal production of MCP-1 in LMVEC was achieved with TNF-α (28.4 ng/ml, P < 0.01), whereas IL-1 was the most potent stimulator of ENA-78 (2.78 ng/ml, P < 0.001) and Groα (29.2 ng/ml, P < 0.001). IL-8 production in LMVEC cells was maximal after LPS stimulation (28.4 ng/ml), but lower than on HUVEC (P < 0.01). LPS, TNF-α and IL-1 stimulation strongly up-regulated all chemokine mRNA. No quantitative differences in mRNA expression between LMVEC and HUVEC were detected for MCP-1 and Groα after LPS stimulation. mRNA expression of ENA-78, GCP-2, CX3C and NAP-2 was however higher in LMVEC under LPS stimulation. In contrast, IL-8 mRNA was slightly more expressed in HUVEC under these conditions. These results further support the hypothesis that the microvascular lung endothelium plays an active role in the induction and perpetuation of acute lung injury.
Antibodies from systemic lupus erythematosus (SLE) sera define differential release of autoantigens from cell lines undergoing apoptosis
Authors: Huggins M L , Todd I , Cavers M A , Pavuluri S R , Tighe P J , Powell R J
Published: March 2003

SLE is an autoimmune disease characterized by a wide range of anti-cellular and anti-nuclear autoantibodies. Many of these antigens are exposed or altered during apoptosis when the nucleus is dismantled in a controlled manner by caspases. We used Western blotting techniques to demonstrate that autoantibodies in SLE sera recognize antigens released during apoptosis. Reproducible bands, not seen in the untreated cells, with the characteristics of histones were seen when staining apoptotic cell lysates with SLE sera. Normal sera recognized some of these bands but much less strongly. Different triggers of apoptosis did not produce marked differences in the antigens recognized. We also compared different cell lines (Jurkat and U937) and found that the staining differed for one autoantigen in particular. The differential release of autoantigens by apoptotic cells may have relevance to the variety of autoantibodies seen in SLE.
Platelet expression of tumour necrosis factor-alpha (TNF-α), TNF receptors and intercellular adhesion molecule-1 (ICAM-1) in patients with proliferative diabetic retinopathy
Authors: Limb G A , Webster L , Soomro H , Janikoun S , Shilling J
Published: March 2003

Microvascular complications of insulin-dependent diabetes mellitus (IDDM) have been strongly associated with platelet abnormalities, whilst TNF-α has been implicated in the pathogenesis of this condition. However, at present it is not clear whether human circulating platelets express TNF-α or TNF receptors (TNF-R) or whether impaired expression of these molecules and of the TNF-reactive adhesion molecule ICAM-1 may be associated with platelet abnormalities in patients with IDDM. On this basis we investigated the platelet expression of these molecules in patients with IDDM complicated or uncomplicated by proliferative diabetic retinopathy (PDR) and in healthy subjects. We observed that the proportion of platelets staining for TNF-α was significantly higher in IDDM patients with active PDR than in patients without microvascular complications (P = 0.0078), quiescent PDR (P = 0.003) or healthy subjects (P = 0.0013). Patients with active PDR also showed a higher proportion of platelets expressing TNF-RI (P = 0.0052) and TNF-RII (P = 0.015) than healthy controls or patients with quiescent PDR (P = 0.009 and 0.0006, respectively). In addition, the percentage of ICAM-1+ platelets was significantly higher in patients with active PDR than in patients with quiescent PDR (P = 0.0065) or normal subjects (P = 0.013). There was a direct correlation between platelet expression of TNF-α and that of TNF-R in PDR patients, indicating that platelet staining for TNF-α may be due to binding of this cytokine to its receptors. The results suggest that increased platelet expression of TNF-α, TNF-R and ICAM-1 in IDDM patients may constitute important markers of thrombocyte abnormalities during the development of microvascular complications of diabetes mellitus.
Expression and production of the long pentraxin PTX3 in rheumatoid arthritis (RA)
Authors: Luchetti M M , Piccinini G , Mantovani A , Peri G , Matteucci C , Pomponio G , Fratini M , Fraticelli P , Sambo P , Di Loreto C , Doni A , Introna M , Gabrielli A
Published: March 2003

PTX3 is a secreted molecule which consists of a C-terminal domain similar to classical pentraxins (e.g. C-reactive protein (CRP)) and of an unrelated N-terminal domain. Unlike the classical pentraxins, the long pentraxin PTX3 is expressed in response to IL-1β and tumour necrosis factor-alpha (TNF-α), but not to IL-6, in various cell types. The present study was designed to investigate the expression of PTX3 in RA. Dissociated RA and osteoarthritis (OA) type B synoviocytes were cultured in the presence and in the absence of inflammatory cytokines. PTX3 mRNA expression in synoviocytes was evaluated by Northern analysis. PTX3 protein levels in synovial cell cultures and synovial fluid were estimated by ELISA, and PTX3 distribution in synovial tissues by immunohistochemical techniques. OA synoviocytes were induced to express high levels of PTX3 mRNA by TNF-α, but not by other cytokines including IL-1β and IL-6. RA synoviocytes, unlike OA synoviocytes, constitutively expressed high levels of PTX3 in the absence of deliberate stimulation. The constitutive expression of PTX3 in RA synoviocytes was not modified by anti-TNF-α antibodies, IL-1 receptor antagonist or a combination of the two agents. In contrast, interferon-gamma and transforming growth factor-beta inhibited PTX3 constitutive expression in RA synoviocytes. The joint fluid from RA patients contained higher levels of immunoreactive PTX3 than controls and the synovial tissue contained endothelial cells and synoviocytes positive for PTX3 by immunohistochemistry. In conclusion, PTX3 may play a role in inflammatory circuits of RA, and its relevance as a marker of disease activity deserves further study.
Cytokines and soluble cytokine receptor induction after IL-12 administration in cancer patients
Authors: Haicheur N , Escudier B , Dorval T , Negrier S , De Mulder P H M , Dupuy J M , Novick D , Guillot T , Wolf S , Pouillart P , Fridman W H , Tartour E
Published: March 2003

This study shows that subcutaneous administration of increasing doses of IL-12, once a week, in 21 cancer patients increased the expression of cytokine genes (interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α), IP-10, MIG, IL-10, IL-4) in peripheral blood mononuclear cells even at very low doses (30 ng/kg). Surprisingly, no circulating TNF-α or IL-4 could be detected in the plasma of patients treated with IL-12. However, a marked increase of soluble IL-4 receptor was demonstrated in the plasma of five of the six patients studied, which may represent an additional mechanism by which IL-12 inhibits the development of the Th2 response in vivo. A marked decline of IFN-γ and IP10 induction was recorded after repeated cycles of IL-12. In contrast, in most patients IL-12 increased IL-10 expression with no subsequent decrease during the course of therapy, and even an earlier peak of IL-10 induction at the 6th cycle. In addition, a constant up-regulation of serum soluble IFN-γ receptor levels was observed after each cycle of IL-12 treatment with a delayed peak compared with the IFN-γ peak. The constant rise of IL-10 and soluble IFN-γ receptor during IL-12 therapy may therefore contribute to the inhibition of IFN-γ activity detected after repeated cycles of IL-12. Lastly, a marked heterogeneity of cytokine induction was observed from one patient to another, which appeared to be independent of the dose of IL-12 administered. These data may lead to a better understanding of the biological activity of IL-12 and the in vivo mechanisms of its regulation.
Exacerbation of acute inflammatory arthritis by the colony-stimulating factors CSF-1 and granulocyte macrophage (GM)-CSF: evidence of macrophage infiltration and local proliferation
Authors: Bischof R J , Zafiropoulos D , Hamilton J A , Campbell I K
Published: March 2003

CSF-1 and GM-CSF have been implicated in the pathogenesis of rheumatoid arthritis. We report the effects of CSF-1 and GM-CSF in the development of an acute methylated bovine serum albumin (mBSA)-induced murine arthritis model. Examination of histopathological features revealed that the systemic administration of CSF-1 or GM-CSF following mBSA administration into the knee resulted in the exacerbation of arthritis. This included synovial hyperplasia and joint inflammation, most evident at 7 and 14 days post-mBSA administration, and the appearance of erosive pannus tissue. The exacerbation by CSF-1 and GM-CSF was not sustained but declined in incidence and severity by 21 days post-mBSA administration, similar to the effects of IL-1β in this model, reported here and previously. Macrophages expressing Mac-2 and F4/80 were a prominent feature of the pathology observed, particularly the infiltration of Mac-2+ macrophages seen in all mice administered CSF-1, GM-CSF or IL-1β. Present in inflamed knees was a locally dividing population of cells which included Mac-2+ and F4/80+ macrophages. These studies demonstrate that CSF-1 and GM-CSF can exacerbate and prolong the histopathology of acute inflammatory arthritis and lend support to monocytes/macrophages being a driving influence in the pathogenesis of inflammatory arthritis.
CD4+ and CD8+ T lymphocyte regeneration after anti-retroviral therapy in HIV-1-infected children and adult patients
Authors: Franco J M , León-Leal J A , Leal M , Cano-Rodriguez A , Pineda J A , Macías J , Rubio A , Rey C , Sanchez B , Lissen E
Published: March 2003

Previous studies have shown a slow recovery of naive CD4+ T cell counts after anti-retroviral therapy in HIV-1-infected adults, which is in accordance with thymus atrophy after puberty. Here we investigate whether or not different patterns of naive CD4+ and CD8+ T cell repopulation are present in adult and child patients undergoing anti-retroviral treatment. Thus, 25 adults under highly active anti-retroviral therapy and 10 children under combined anti-retroviral therapy were retrospectively analysed for T cell subpopulations at baseline (T0) and around week 12 (T1) and week 24 (T2) of anti-retroviral treatment. Mean serum HIV-1 RNA levels dropped in both groups. Recovery of T cells in adults was characterized by a heterogeneous response between patients, with only 44% of them increasing their naive CD4+ and CD8+ T cell counts at T1, and changes in mean total CD4+ T cells were mainly shaped by memory cells. Otherwise, children were characterized by an early increase in naive T cells. Thus, at T1, all children analysed had a strong rise in CD4+ (from 389 ± 116 to 569 ± 121 cells/μl; P < 0·01), and nine out of 10 also in naive CD8+ T cells (from 244 ± 58 to 473 ± 85 cells/μl; P < 0·05). However, no significant correlation between age and naive repopulation was observed (P= 0·22) in children. Thus, children had the earlier and greater increases in naive T cell subsets than adults, probably due to a more active thymus, with the potential for immune reconstitution when HIV-1 replication is controlled.
Cellular localization of complement C3 and C4 transcripts in intestinal specimens from patients with Crohn’s disease
Authors: Laufer J , Oren R , Goldberg I , Horwitz A , Kopolovic J , Chowers Y , Passwell J H
Published: March 2003

It has been suggested that the increase in C3 and C4 levels in jejunal perfusates of patients with Crohn’s disease (CD) results from local intestinal synthesis of complement. The present study evaluated the expression of these complement genes in inflamed tissues from patients with CD. Surgically resected specimens from patients with CD and control tissue obtained from subjects with adenocarcinoma of the colon were evaluated for C3 and C4 gene expression by the use of 35S-labelled anti-sense RNA probes. All tissue samples, diseased and normal tissue, expressed C4 mRNA throughout in the intestinal epithelium. C3 mRNA was not detected in epithelial cells in histologically normal tissue, but in diseased specimens there was a focal distribution of C3 mRNA in epithelial cells of the crypts, but not in villous epithelium. Focal C3 gene expression correlated with crypt abscess formation and the presence of polymorphonuclear leucocytes in the lumen of the crypts. In addition, C3 mRNA was also found in macrophages of the submucosa. These macrophages were CD68+, fusiform with faint cytoplasm and morphologically different from the large rounded lamina propria macrophages, which do not express C3 mRNA. Multinucleated giant cells did not express either C3 or C4 genes. In addition to its presence in intestinal epithelium, C4 mRNA was also expressed in mast cells, which however did not express C3 mRNA. These observations identify cells in the intestinal wall expressing complement genes and support the hypothesis that there is local regulated production of complement in the intestine of patients with CD, and subsequent complement activation may contribute to the inflammatory process.
Complex pattern of Th1 and Th2 activation with a preferential increase of autoreactive Th1 cells in BALB/c mice with proteoglycan (aggrecan)-induced arthritis
Authors: Holló K , Glant T T , Garzó M , Finnegan A , Mikecz K , Buzás E
Published: March 2003

The central role of CD4+ T cells and the balance between T helper (Th) subpopulations in the pathogenesis of autoimmune diseases have been extensively studied. Proteoglycan (aggrecan)-induced arthritis (PGIA) is a murine model for rheumatoid arthritis (RA), which is characterized by a Th1 dominance at the onset of the disease. In addition to CD4+ T cells, antigen-presenting B cells and autoantibodies seem to play an important role in the development and regulation of PGIA. To identify proteoglycan-specific CD4+ T cell subsets and Th1- and Th2-supported antibody isotypes during the progression of PGIA, spleen cells of proteoglycan-immunized BALB/c mice were harvested at different times of immunization, and at different stages of the disease, and their cytokine production and antigen-specific antibody isotype profiles were determined by enzyme-linked immunospot (ELISPOT) assays. Both Th1 and Th2 cytokine-producing cells, with the predominance of IL-4/IL-5-secreting cells, were detected during the prearthritic stage, and a shift toward a Th1 dominance was observed at the time of onset of arthritis. Tissue homogenates of acutely inflamed joints contained significantly higher levels of interferon-gamma than IL-4. The prearthritic period and both the acute and chronic phases of joint inflammation were characterized by IgG1 dominance in the sera and this correlated with the number of IgG1-secreting B cells in the spleen. However, the ratio of autoreactive IgG1/IgG2a-secreting cells decreased in arthritic animals. These results indicate the activation and possible regulatory roles of both Th1 and Th2 subsets in the autoimmune process, with the necessity of a relative increase of autoreactive Th1 cells for the induction of joint inflammation.
Association of neutropenia in systemic lupus erythematosus (SLE) with anti-Ro and binding of an immunologically cross-reactive neutrophil membrane antigen
Authors: Kurien B T , Newland J , Paczkowski C , Moore K L , Scofield R H
Published: March 2003

SLE is associated with the production of autoantibodies to self-constituents. In particular, certain ribonucleoprotein particles are targeted. Despite the multitude of autoantibodies produced and the remarkable concentrations of these antibodies in the sera of SLE patients, there have been little data that the autoantibodies found in SLE are involved in the pathogenesis of disease or its manifestations. The present work demonstrates that anti-Ro (or SSA) is associated with granulocytopenia, binds the surface of granulocytes and fixes complement to this membrane surface. Binding is a property of anti-Ro Fab fragments and can be inhibited by 60-kD Ro. However, the antigen bound on the surface of granulocytes is a 64 000 mol. wt protein that is a novel autoantigen in SLE. As suggested by inhibition studies, sequence identity between 60-kD Ro and eight tandem repeats in the 64-kD antigen may be responsible for the observed serologic cross-reactivity . These data imply that anti-Ro antibodies that also bind the 64-kD protein mediate neutropenia in patients with SLE.
IL-17 regulates gene expression and protein synthesis of the complement system, C3 and factor B, in skin fibroblasts
Authors: Katz Y , Nadiv O , Rapoport M J , Loos M
Published: March 2003

Human IL-17 is a cytokine secreted by CD4+-activated memory T cells with the profile of effects of a Th1 cytokine. The effects of IL-17 on many cellular constituents of joints suggest that it may participate in inflammatory joint diseases. Proteins of the complement system are known to be regulated by pro- and anti-inflammatory cytokines. The purpose of this work was to study the effect of IL-17 alone and combined with tumour necrosis factor (TNF) on the expression and synthesis of factor B and C3. Fibroblasts were stimulated with the relevant cytokine or cytokines, pulse labelled with 35S-methionine, and the newly synthesized proteins were immunoprecipitated and subjected to SDS–PAGE. Gene expression was determined by Northern blot analysis. IL-17 10 ng/ml induced increases in gene expression and protein synthesis of C3, 2·25 ± 0·26- and 2·7 ± 0·7-fold, respectively, with concomitant non-significant effects on factor B, 1·5 ± 0·45- and 2·2 ± 1·2-fold, respectively. When both IL-17 and TNF were present simultaneously, the synthesis of factor B increased by 85% more than the expected additive effects of these cytokines separately, while for C3 the effect of both cytokines was 19% lower than the expected additive effect (observed/expected = 0·81). IL-4 reduced the synergistic effect by 50%. We conclude that IL-17 has a regulatory role on C3 expression and synthesis and an amplifying effect on TNF-induced factor B synthesis. Taken together with the evidence that TNF is a major cytokine involved in the inflammation of rheumatoid arthritis, it suggests that IL-17 has a proinflammatory role in the inflammation process of joints. The distinct effects of IL-4, IL-17 and TNF on the synthesis of factor B in fibroblasts suggest that factor B and the alternative pathway of the complement system may play an important role in joint inflammation.
Cytolytic mechanisms of intraepithelial lymphocytes in coeliac disease (CoD)
Authors: Ciccocioppo R , Di Sabatino A , Parroni R , D’Alò S , Pistoia M A , Doglioni C , Cifone M G , Corazza G R
Published: March 2003

The effector arm of the mucosal immune system comprises lymphocytes scattered at intraepithelial and lamina propria levels. Intraepithelial lymphocytes (IEL) are a large population of oligoclonal resting cells which exhibit phenotypic and functional characteristics of cytolytic T cells when activated. Several mechanisms have been demonstrated to account for their cytotoxicity. Among them, one is mediated by perforin and granzyme molecules, another is mediated by Fas ligand (FasL) which delivers apoptotic signals through Fas receptor on target cells. There is good evidence that a flat intestinal mucosa may be produced by activated T cells. The aim of our study was to evaluate FasL and perforin expression by IEL, and its possible correlation with the increased enterocyte apoptosis in coeliac mucosa. Endoscopic duodenal biopsy specimens from 10 untreated coeliac patients, 10 treated coeliac patients, and 10 biopsied controls were evaluated for enterocyte apoptosis by terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine triphosphate nick end label method, for perforin expression by immunohistochemistry, and for FasL expression by immunocytochemistry. In untreated CoD there was a significant increase of percentage of both FasL+ and perforin+ IEL which positively correlated with enterocyte apoptosis in comparison with controls. All these parameters were significantly lower in treated CoD, even though they did not normalize. Our study demonstrates that in untreated CoD FasL and perforin expression by IEL is increased, and significantly correlates with the level of enterocyte apoptosis.
Decreased CD95 expression on naive T cells from HIV-infected persons undergoing highly active anti-retroviral therapy (HAART) and the influence of IL-2 low dose administration
Authors: Amendola A , Poccia F , Martini F , Gioia C , Galati V , Pierdominici M , Marziali M , Pandolfi F , Colizzi V , Piacentini M , Girardi E , D’Offizi G
Published: March 2003

The functional recovery of the immune system in HIV-infected persons receiving HAART and the role of adjuvant immune therapy are still matters of intensive investigation. We analysed the effects of HAART combined with cytokines in 22 naive asymptomatic individuals, randomized to receive HAART (n = 6), HAART plus a low dose (1000 000 U/daily) of rIL-2 (n = 8), and HAART plus rIL-2 after previous administration of granulocyte colony-stimulating factor (n = 8). After 3 months of therapy, increased CD4+ T cell counts and diminished viral loads were observed in all patients, independently of cytokine addition. A decreased expression of CD95 (Apo 1/Fas) was evident in all groups when compared with values before therapy. The percentages of peripheral blood mononuclear cells (PBMC) expressing CD95 after therapy decreased by 15%, 22% and 18% in the three treatment groups, respectively (P < 0·05). Analysis of PBMC subsets demonstrated that CD95 expression was significantly reduced on CD45RA+CD62L+ naive T cells (25·3%, 22·4%, and 18·6%, respectively; P < 0·05) in each group, after therapy. Accordingly, all patients showed a reduced rate of in vitro spontaneous apoptosis (P < 0·05). Another effect induced by HAART was a significant increase in IL-2Rα expression on total PBMC (P < 0·05), independently of cytokine addition. Altogether, our results suggest that very low dose administration of rIL-2 (1000 000 U/daily) may be not enough to induce a significant improvement in the immune system as regards HAART alone. The employment of higher doses of recombinant cytokines and/or different administration protocols in clinical trials might however contribute to ameliorate the immune reconstitution in patients undergoing HAART.
Co-stimulatory signals increase the reactivity ofγδT cells towards mycobacterial antigens
Authors: González-Amaro R , Portales-Pérez D P , Baranda L , Moncada B , Toro C , López-Briones S , Espitia C , Mancilla R
Published: March 2003

Although it has been shown that γδ T lymphocytes are able to react with different cell-associated or soluble antigens, the immune repertoire of these cells appears to be skewed to the recognition of mycobacterial antigens. We have studied the number and reactivity of γδ T cells towards several mycobacterial antigens in patients with tuberculosis and leprosy, as well as their healthy contacts and control individuals. We found an increased number of Vδ2+ cells in healthy contacts (PPD+ and lepromin+) and tuberculoid leprosy patients. The γδ T cells from lepromatous leprosy showed a decreased response to all antigens tested, but some of these patients exhibited a significant response to the 30-kD glycoprotein of Mycobacterium tuberculosis. Interestingly, the reactivity of γδ T cells against mycobacterial antigens was significantly increased by costimulatory signals generated through CD7, LFA-1, CD50 and CD69 in all groups. However, signalling through CD69 did not enhance the responsiveness of γδ lymphocytes from lepromatous patients. On the other hand, the in vitro blockade of IL-10 with a specific antibody enhanced the cell proliferation of γδ lymphocytes from lepromatous leprosy patients, whereas exogenous IL-10 had an opposite effect in most individuals studied. These results suggest the potential role of different cell membrane receptors in the regulation of γδ T cell proliferation induced by mycobacteria, as well as the possible involvement of IL-10 in this phenomenon.
Inhibition of cyclin A gene expression in human B cells by an immunosuppressant mizoribine
Authors: Hirohata S , Nakanishi K , Yanagida T
Published: March 2003

Mizoribine has been shown to have beneficial effects in the treatment of rheumatoid arthritis and lupus nephritis, in which abnormal B cell functions are involved. Previous studies demonstrated that mizoribine directly suppresses the function of human B cells. The current study explored in detail the mechanism of the suppression of human B cell responses by mizoribine at the molecular level. Highly purified peripheral blood B cells obtained from normal healthy individuals were stimulated with Staphylococcus aureus Cowan I (SAC) plus IL-2 in the presence or absence of mizoribine or methotrexate for 48 h to 72 h. The expression of cyclin A mRNA was determined by semiquantitative reverse transcriptase-polymerase chain reaction followed by Southern hybridization. Although at pharmacologically attainable concentrations both mizoribine and methotrexate suppressed the production of IgM of SAC-activated B cells, mizoribine, but not methotrexate, decreased the expression of cyclin A protein as well as mRNA in B cells stimulated with SAC + IL-2. Of note, mizoribine facilitated the degradation of cyclin A mRNA in the presence of actinomycin D, indicating that mizoribine shortens the stability of cyclin A mRNA. The results indicate that mizoribine suppresses the expression of cyclin A mRNA in human B cells by down-regulating its stability, and thus down-regulates their responses.
Cyclosporin A reduces expression of adhesion molecules in the kidney of rats with chronic serum sickness
Authors: Rincón J , Parra G , Quiroz Y , Benatuil L , Rodríguez-Iturbe B
Published: March 2003

Treatment with cyclosporin A (CsA) improves proteinuria and reduces renal cellular infiltration in chronic serum sickness (CSS). We examined if these effects were associated with a reduced renal expression of CD54 and its ligands, interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α) and MHC class II molecules. We studied two groups of rats in which CSS was induced by daily injections of ovalbumin (OVA): a group treated with CsA (OVA.CsA group, n = 11) and a group that received no treatment (OVA.CSS group, n = 11). An additional group of five rats (control group) received only phosphate buffer. Immunostaining techniques were used to follow CSS and to study the expression of CD54, CD18, CD11b/c, IFN-γ, TNF-α and MHC class molecules. Proteinuria (mg/24 h) was reduced from 248.2 ± 73.1 (OVA.CCS group) to 14.5 ± 13.1 with CsA treatment (P < 0.0001). The renal expression of CD54 and its ligands (CD18 and CD11b/c) was reduced by 50% to 75%. Correspondingly, there was a 60% to 85% reduction in the number of infiltrating leucocytes. The number of cells expressing TNF-α, IFN-γ and MHC II molecules was also reduced. CsA reduces expression of CD54 and its ligands. This effect is associated with a reduction of cellular infiltration, IFN-γ, TNF-α-producing cells and with MHC II expression in the kidney. These findings suggest that expression of adhesion molecules plays a critical role in CSS and underline the importance of cellular immunity in this experimental model.
Staphylococcal toxin-induced T cell proliferation in atopic eczema correlates with increased use of superantigen-reactive Vβ-chains in cutaneous lymphocyte-associated antigen (CLA)-positive lymphocytes
Authors: Davison S , Allen M , Vaughan R , Barker J
Published: March 2003

Staphylococcal superantigens have been implicated in the pathogenesis of atopic dermatitis (AD). This may occur through superantigenic activation of T lymphocytes and their subsequent induction of the skin homing receptor CLA on activated cells. We investigated the proliferative responses of peripheral blood mononuclear cells (PBMC) from 10 patients with an infective exacerbation of AD and six normal controls to the staphylococcal superantigens, staphylococcal enterotoxin A and B (SEA, SEB) and toxic shock syndrome toxin-1 (TSST-1), and the mitogens phytohaemagglutinin (PHA) and concanavalin A (Con A). We also assessed CLA and T cell receptor (TCR) Vβ-chain expression by immunofluorescence and flow cytometry before and after stimulation. PBMC from AD patients showed two-fold increased proliferation to SEA and SEB (P < 0·01) compared with normals, whereas the response to mitogenic stimulation was identical. Analysis of (TCR) Vβ-chain expression demonstrated increased use of superantigen-reactive Vβ families in freshly isolated PBMC in AD patients compared with controls. This pattern of Vβ-chain expression was only observed in the CLA+ but not the total population of T cells. Furthermore, there was a positive correlation between the enhanced PBMC proliferative response and increased expression of superantigen-reactive Vβ families in atopic patients. These data support the concept that superantigens are important in the pathogenesis of this common condition, and also provide evidence that the increased use of certain Vβ families in circulating, CLA+, skin homing lymphocytes is of functional significance.
Expression of vascular endothelial growth factor by synovial fluid neutrophils in rheumatoid arthritis (RA)
Authors: Kasama T , Kobayashi K , Yajima N , Shiozawa F , Yoda Y , Takeuchi H T , Mori Y , Negishi M , Ide H , Adachi M
Published: March 2003

Most of the leucocytes infiltrating rheumatoid synovial fluid (SF) are neutrophils capable of producing a variety of inflammatory mediators known to contribute significantly to the disease process during active RA. In the present study, we investigated the contribution made by SF neutrophils to the elevated levels of vascular endothelial growth factor (VEGF) seen in rheumatoid SF. Rheumatoid SF neutrophils were found to contain significantly larger amounts of both VEGF protein and its mRNA than peripheral blood neutrophils from either RA patients or healthy controls. Levels of celL-associated VEGF were well correlated with free VEGF in SF, which was significantly higher than in SF from osteoarthritis patients. Levels of SF neutrophiL-associated VEGF also correlated with RA disease activity and cell surface integrin expression. Thus, SF neutrophiL-associated VEGF may be considered an indicator of both local and systemic inflammation of RA, contributing to the neovascularization seen during RA synovitis.
Booster immunization of children with an acellular pertussis vaccine enhances Th2 cytokine production and serum IgE responses against pertussis toxin but not against common allergens
Authors: Ryan E J , Nilsson L , Kjellman N-I M , Gothefors L , Mills K H G
Published: March 2003

Acellular pertussis vaccines (Pa) protect against severe pertussis in children. However, serum antibody responses decline quickly after immunization. Studies in animal models suggest that cell-mediated immunity also contributes to protection against Bordetella pertussis, and it has already been demonstrated that Pa induce T cells that secrete type-1 and type-2 cytokines in children. In this study we examined the persistence of the T cell response and the effect of booster immunization in 4–6-year-old children. Cell-mediated immunity to B. pertussis antigens was detected in a high proportion of children more than 42 months after their last immunization. Peripheral blood mononuclear cells (PBMC) from the majority of children secreted interferon-gamma (IFN-γ) and a smaller proportion IL-5, in response to specific antigen stimulation in vitro. However, following booster immunization, significantly higher concentrations of IL-5, but not IFN-γ, were produced by PBMC in response to B. pertussis antigens. Furthermore, plasma IL-4 and IL-5 concentrations were increased, whereas IFN-γ concentrations were reduced following booster immunization. It has been suggested that childhood immunization with Th2-inducing vaccines may predispose some children to atopic disease. Although we found that pertussis toxin (PT)-specific IgE was significantly increased after booster immunization in both atopic and non-atopic children, the levels of IgE to common allergens and the prevalence of positive skin prick test were unaffected by the booster vaccination. Thus, despite the enhancement of type-2 responses to B. pertussis antigens, booster vaccination with Pa does not appear to be a risk factor for allergy.
B cell epitopes of gliadin
Authors: Osman A A , Günnel T , Dietl A , Uhlig H H , Amin M , Fleckenstein B , Richter T , Mothes T
Published: March 2003

A phage displayed dodecapeptide library and synthetic octapeptides spanning the complete sequence of α- and γ-type gliadin and overlapping in six amino acids (pepscan) were screened for binding to human gliadin antibodies (AGA). Phage display experiments led to four sequences recognized with significantly higher frequency by sera with raised IgA-AGA titres than by control sera. All these peptides contained the core sequence PEQ. Pepscan experiments revealed binding of AGA to five prominent regions: (i) QXQPFP (binding to IgG and IgA, X representing P, Q, and L); (ii) IPEQ (IgG) and WQIPEQ (IgA); (iii) FFQP (IgG) and QGXFQP (IgA, X representing F and S); (iv) PQQLPQ (IgG and IgA), all in α-type gliadin; and (v) QPQQPF (IgG and IgA) in γ-type gliadin. In two of the sequences (QPQQPF and QQQPFP), substitution of Q by E resulting in QPEQPF and QEQPFP, respectively, increased significantly binding of AGA from sera of patients with biopsy-proven or suspected coeliac disease (CoD), all positive for endomysium antibodies (EmA). In contrast, binding of sera with high AGA titre from EmA-negative patients (CoD and dermatitis herpetiformis excluded) was not enhanced by this substitution. Thus, AGA directed against these modified epitopes can be regarded as specific for CoD. This is the first study demonstrating that deamidation of gliadin improves reactivity of AGA of CoD patients.
Significance of chemokines and activated platelets in patients with diabetes
Authors: Nomura S , Shouzu A , Omoto S , Nishikawa M , Fukuhara S
Published: March 2003

Levels of platelet-derived microparticles (PMPs), platelet activation markers (P-selectin expressed on, or annexin V binding to, platelets (plt:P-selectin or plt:annexin V, respectively)), chemokines (IL-8, monocyte chemotactic peptide-1 (MCP-1), and regulated on activation normally T-cell expressed and secreted (RANTES)), and soluble P- and E-selectins were compared in peripheral blood from diabetic and control patients in order to develop a better understanding of their potential contribution to diabetic vascular complications. Significant increases were found for PMPs, plt:P-selectin, MCP-1, RANTES and soluble P- and E-selectins in diabetic individuals, whereas IL-8 levels were similar. Furthermore, after ticlopidine treatment, most of these factors receded to baseline levels observed in non-diabetic patients. Our findings indicate that ticlopidine might be able to prevent or reduce vascular complications in diabetic patients.