bluuebunny/crossref_metadata_2025 huggingface dataset:
Pigeon fanciers' lung: effects of smoking on serum and salivary antibody responses to pigeon antigens
Authors: Baldwin C I , Todd A , Bourke S , Allen A , Calvert J E
Published: March 2003

A reduced prevalence of pigeon fanciers' lung has been reported in pigeon breeders who smoke cigarettes. Serum and salivary antibodies to pigeon intestinal mucin and pigeon serum proteins were investigated in 227 pigeon fanciers, subdivided according to smoking habit and clinical status. Smokers had a lower incidence of precipitating antibodies to pigeon antigens and lower titres of serum IgG and IgA antibodies to mucin and to pigeon serum proteins in ELISA compared with non-smokers and ex-smokers. In contrast, IgG antibody titres to tetanus toxoid were similar in smoking and non-smoking groups. In contrast to serum antibodies, salivary IgA antibody titres to pigeon antigens were similar in smokers and non- or ex-smokers. Approximately one third of the smokers reported symptoms consistent with pigeon fanciers' lung but did not have precipitating antibodies. Only some individuals with precipitating antibodies had disease symptoms, and IgG antibody titres in these individuals were not significantly higher than in many asymptomatic individuals. Salivary IgA titres against pigeon mucin were significantly higher in asymptomatic individuals, consistent with a protective role for these antibodies. The results confirm that smoking is associated with a decreased serum antibody response to inhaled pigeon antigens, affecting IgG1, IgG2 and IgA responses, but this impairment does not extend to salivary IgA or to antibody responses to a parenterally administered protein antigen. The fact that responses to pigeon serum proteins and to pigeon intestinal mucin were similarly affected suggests that cigarette smoking depresses both T-independent and T-dependent responses to inhaled antigens.
Origin and properties of soluble CD21 (CR2) in human blood
Authors: Ling N R , Hardie D L , Johnson G D , Maclennan I C M
Published: March 2003

By analysis with a panel of CD21 MoAbs it is shown that a large part of the soluble CD21 in human blood plasma is of the long isoform (CD21L), as judged by comparison with antigen produced by mouse L cells transfected with CD21L-cDNA and reactivity with the restricted CD21 MoAb R4/23. This is compatible with the hypothesis that soluble CD21 in the blood is mainly derived from follicular dendritic cells (FDC). Cells from a human keratinocyte cell line transfected with cDNA from the Burkitt lymphoma cell line Raji also produced soluble CD21L (sCD21L), whereas the short form of sCD21 (sCD21S) was the major component of sCD21 produced by the B lymphoblastoid cell line LICR-LON-HMy and the T cell line Jurkat. Confocal studies of FDC isolated from human tonsil revealed that CD21 was present in the cytoplasm. On gel filtration sCD21 from untreated serum has an apparent size considerably greater than the 130 kD found by SDS–PAGE analysis. This may be partly accounted for by the non-globular shape of the molecule, but may also indicate, as reported by others, that in its native state sCD21 is complexed with other proteins. However, no evidence of complexing with sCD23 or C3d could be found.
Macrophage function in alloxan diabetic mice: expression of adhesion molecules, generation of monokines and oxygen and NO radicals
Authors: Ptak W , Klimek M , Bryniarski K , Ptak M , Majcher P
Published: March 2003

The increased incidence of bacterial and mycotic infections in poorly controlled diabetic patients or animals is frequently attributed to impaired activities of professional phagocytes (granulocytes, macrophages) in hypoinsulinaemic milieu. We measured production of monokines (IL-6 and tumour necrosis factor-alpha (TNF-α)), active NO and reactive oxygen intermediates (ROIs), as well as expression of several cell surface adhesion molecules (Mac-1, -2 and -3, intercellular adhesion molecule-1 (ICAM-1) and FcγRII), by thioglycollate medium-induced peritoneal macrophages of normoglycaemic and alloxan diabetic CBA/J mice (blood glucose level in the range 300 or 500 mg/dl). Macrophages of animals with moderate diabetes (300 mg/dl) produced significantly more IL-6 and TNF-α and ROIs than cells of control mice and showed an increased expression of all cell surface molecules, except Mac-3. NO/NO2 production was not affected. Administration of insulin restored enhanced values to normal levels, except for the production of ROIs which remained unusually high. We conclude that two separate mechanisms influence macrophage physiology in diabetes—lack of saturation of insulin receptors on macrophages and an indirect effect due to formation of advanced glycosylation endproducts (AGE) on their surfaces. The latter is possibly responsible for increased generation of ROIs, since it cannot be down-regulated by prolonged insulin treatment. How the increased activity of macrophages of moderately diabetic mice (enhanced production of proinflammatory monokines and oxygen radicals as well as expression of molecules) is related to their ability to kill bacteria is now under investigation.
Levels of soluble FcγRIII correlate with disease severity in sepsis
Authors: Muller Kobold A C , Zijlstra J G , Koene H R , DE Haas M , Kallenberg C G M , Cohen Tervaert J W
Published: March 2003

Neutrophil activation is thought to play a crucial role in the pathogenesis of sepsis. During activation, neutrophils adhere to and migrate through the endothelium. Therefore, the amount of circulating neutrophils does not adequately reflect the total amount of neutrophils that are involved in the pathophysiologic process of this condition. In this study we test the hypothesis that the severity of sepsis is associated with the total body mass of neutrophils as reflected in the plasma concentration of soluble Fcγ receptor type III (sFcγRIII). Nineteen patients with sepsis (12 male, seven female, median age of 69 years, range 29–87 years) were included in this study. Ten healthy volunteers served as controls. Plasma sFcγRIII concentrations were measured by ELISA. Other parameters that were studied were leucocyte count, plasma concentrations of lactoferrin and soluble l-selectin, and surface expression of CD11b and CD66b on circulating neutrophils. Disease activity was measured using the Acute Physiology and Chronic Health Evaluation (APACHE) II score. Soluble FcγRIII levels were elevated in sepsis patients whereas soluble l-selectin levels were moderately decreased compared with healthy controls. Markers of cell activation were significantly increased in sepsis patients. Soluble FcγRIII correlated with disease severity as measured by the APACHE score (P < 0.05, r = 0.53), whereas the other parameters did not correlate with the APACHE score. In conclusion, this study demonstrates that soluble FcγRIII is a useful marker for disease severity in patients with sepsis.
A mixed population of immature and mature leucocytes in umbilical cord blood results in a reduced expression and function of CR3 (CD11b/CD18)
Authors: Reddy R K , Xia Y , Hanikýřová M , Ross G D
Published: March 2003

Neonatal neutrophils express less membrane and cytoplasmic CR3 (iC3b-receptor, Mac-1, αMβ2-integrin) than do adult neutrophils, and it has been suggested that this renders neonatal neutrophils deficient in diapedesis and bactericidal activity. The reason(s) for this deficiency are unknown. In this study, CR3 expression and the CR3-dependent respiratory burst activity of individual neonatal neutrophils are quantified in comparison with adult leucocytes using flow cytometry. Monocytes and neutrophils are defined as CD14highCD15low and CD14lowCD15high, respectively. Although neonatal neutrophils bore less CR3 on average than did adult neutrophils, neonatal neutrophils were more heterogeneous and many neonatal neutrophils expressed adult levels of CR3. Because of higher neutrophil concentrations in cord versus adult blood, the calculated number of neutrophils in cord blood expressing high amounts of CR3 was equivalent to that of adult blood. Similar findings were made with monocytes. The size of the CR3-dependent respiratory burst stimulated by particulate β-glucan correlated directly with the expression of CR3 by individual neutrophils. With neonatal and adult neutrophils having comparable CR3 densities, the respiratory burst activities were equivalent. Wright–Giemsa differential staining of the subset of neonatal neutrophils with low CR3 levels isolated by fluorescence-activated cell sorting showed a higher proportion of immature cells than the sorted population expressing high CR3 levels. Therefore, higher proportions of immature cells in cord blood probably explain previous reports of deficient CR3 expression and function. The typical neutrophilia of cord blood may compensate for this apparent deficiency by providing adult concentrations of mature neutrophils.
Reduced production of both Th1 and Tc1 lymphocyte subsets in atopic dermatitis (AD)
Authors: LONATI A , LICENZIATI S , CANARIS A D , FIORENTINI S , PASOLINI G , MARCELLI M , SEIDENARI S , CARUSO A , DE PANFILIS G
Published: March 2003

An imbalance of interferon-gamma (IFN-γ)-bearing CD4+ T (Th1) cells in the pathogenesis of AD is well recognized; however, a possible role in AD for CD8+ T cells secreting Th1-like cytokines (Tc1) has not been properly addressed. In this study, two- and three-colour FACS analysis allowed us to discriminate the Th1 from the Tc1 subset. AD patients had half the number of IFN-γ-producing circulating T cells (P < 0.005; 13.6 ± 1.9% (mean ± s.d.)) compared with normal donors (25.0 ± 2.4%). Specifically, both Th1 (4.8 ± 0.7%) and Tc1 (8.1 ± 1.1%) cells in AD were decreased compared with Th1 (8.8 ± 0.8%) and Tc1 (15.0 ± 1.5%) cells in controls. Moreover, at the mRNA level, the ratios of IFN-γ/IL-4 and IFN-γ/IL-10 were lower in cells from AD patients compared with controls. In conclusion, the decrease of IFN-γ-producing T lymphocytes in AD is due to a reduction in both Th1 and Tc1 IFN-γ-secreting cells; this may not only contribute to the over-production of IgE, but also explain the high incidence of cutaneous infections observed in AD patients.
Antigenicity of theLeishmania infantumhistones H2B and H4 during canine viscerocutaneous leishmaniasis
Authors: SOTO M , REQUENA J M , QUIJADA L , PEREZ M J , NIETO C G , GUZMAN F , PATARROYO M E , ALONSO C
Published: March 2003

In this study we show that sera from dogs naturally infected with Leishmania infantum contain antibodies that specifically react against the parasite H2B and H4 histones. The Leishmania H2B and the amino-terminal region of the histone H4, expressed as fusion proteins, when confronted with sera from canine viscerocutaneous leishmaniasis (VCL) dogs, were recognized by 63% and 47%, respectively. No reactivity was detected when sera from dogs naturally infected with pathogens other than Leishmania were used. Using a collection of synthetic peptides covering the complete sequence of both proteins, we have determined that the main linear antigenic determinants are located in the amino-terminal domains of these histones. The humoral response against histones H2B and H4 induced during canine leishmaniasis was found to be specific for Leishmania histones, since no cross-reactivity of the VCL sera with mammal histones was observed. Also, a comparative study of the prevalence of antibodies among VCL sera against the four core histones of L. infantum was performed. Although a large heterogeneity of the humoral responses against these proteins was found, histones H2A and H3 seem to be more prevalent immunogens than histones H2B and H4 during canine natural leishmaniasis. The origin of the anti-histone humoral response and its possible implications in the pathogenesis of Leishmania infection are discussed.
HBV-specific immune defect in chronic hepatitis B (CHB) is correlated with a dysregulation of pro- and anti-inflammatory cytokines
Authors: SCHLAAK J F , TULLY G , LÖHR H F , GERKEN G , MEYER ZUM BÜSCHENFELDE K-H
Published: March 2003

The aim of this study was to examine the immunomodulating effects of rhIL-12 on the immune response induced by hepatitis B virus (HBV) antigens in clinical subgroups of patients with HBV infection. Peripheral blood mononuclear cells (PBMC) of 80 patients were stimulated with HBsAg, HBcAg, pre-S1Ag and tetanus toxoid in the absence or presence of IL-12 (0.01, 0.1 and 1 ng/ml). Stimulation by anti-CD3 + anti-CD28 and lipopolysaccharide (LPS) were used as controls. Proliferation and cytokine production were determined by 3H-thymidine uptake and ELISA after 72 h. After stimulation with HBV antigens only, production of tumour necrosis factor-alpha (TNF-α) or IL-10 was observed in all patients, while interferon-gamma (IFN-γ) was detectable in only 27 patients. After costimulation with IL-12 and HBV antigens, however, large amounts of IFN-γ were found in all patients, while HBV-induced IL-10 production remained mostly unchanged. When clinical subgroups including patients with compensated liver cirrhosis were compared, PBMC from patients with HBeAg+ hepatitis showed the lowest capacity to produce IFN-γ after HBV antigen-positive IL-12. These data suggest that the ability of IL-12 to enhance IFN-γ production against HBV antigens is correlated with the presence of HBeAg and is not impaired in patients with advanced liver disease. In addition, IL-12 and IL-10 production by antigen-presenting cells may be a critical factor that determines the efficacy of the immune response against the hepatitis B virus.
Anti-mitochondrial flavoprotein autoantibodies of patients with myocarditis and dilated cardiomyopathy (anti-M7): interaction with flavin-carrying proteins, effect of vitamin B2 and epitope mapping
Authors: STÄHLE I , BRIZZIO C , BARILE M , BRANDSCH R
Published: March 2003

Vitamin B2 and flavin cofactors are transported tightly bound to immunoglobulin in human serum. We reasoned that anti-mitochondrial flavoprotein autoantibodies (αFp-AB) present in the serum of patients with myocarditis and cardiomyopathy of unknown aetiology may form immunoglobulin aggregates with these serum proteins. However, immunodiffusion and Western blot assays demonstrated that the flavin-carrying proteins were not recognized by αFp-AB. Apparently the flavin moiety in the native protein conformation was inaccessible to αFp-AB. This conclusion was supported by the absence of an immunoreaction between the riboflavin-binding protein from egg white and αFP-AB. Intravenous application of vitamin B2 to rabbits immunized with 6-hydroxy- d-nicotine oxidase, a bacterial protein carrying covalently attached FAD, did not neutralize αFp-AB which had been raised in the serum of the animals. FAD-carrying peptides generated from 6-hydroxy-d-nicotine oxidase by trypsin and chymotrypsin treatment were not recognized by the αFp-AB, but those generated by endopeptidase Lys were. This demonstrates that the epitope recognized by αFp-AB comprises, besides the flavin moiety, protein secondary structure elements.
An IgG1 titre to the F1 and V antigens correlates with protection against plague in the mouse model
Authors: WILLIAMSON E D , VESEY P M , GILLHESPY K J , ELEY S M , GREEN M , TITBALL R W
Published: March 2003

The objective of this study was to identify an immunological correlate of protection for a two-component subunit vaccine for plague, using a mouse model. The components of the vaccine are the F1 and V antigens of the plague-causing organism, Yersinia pestis, which are coadsorbed to alhydrogel and administered intramuscularly. The optimum molar ratio of the subunits was determined by keeping the dose-level of either subunit constant whilst varying the other and observing the effect on specific antibody titre. A two-fold molar excess of F1 to V, achieved by immunizing with 10 μg of each antigen, resulted in optimum antibody titres. The dose of vaccine required to protect against an upper and lower subcutaneous challenge with Y. pestis was determined by administering doses in the range 10 μg F1 + 10 μg V to 0.01 μg F1 + 0.01 μg V in a two-dose regimen. For animals immunized at the 1-μg dose level or higher with F1 + V, an increase in specific IgG1 titre was observed over the 8 months post-boost and they were fully protected against a subcutaneous challenge with 105 colony-forming units (CFU) virulent Y. pestis at this time point. However, immunization with 5 μg or more of each subunit was required to achieve protection against challenge with 107 CFU Y. pestis. A new finding of this study is that the combined titre of the IgG1 subclass, developed to F1 plus V, correlated significantly (P < 0.05) with protection. The titres of IgG1 in vaccinated mice which correlated with 90%, 50% and 10% protection have been determined and provide a useful model to predict vaccine efficacy in man.
Antibody-independent classical complement pathway activation and homologous C3 deposition in xeroderma pigmentosum cell lines
Authors: KURITA M , MATSUMOTO M , TSUJI S , KAWAKAMI M , SUZUKI Y , HAYASHI H , TOYOSHIMA K , SEYA T
Published: March 2003

Of human malignantly transformed cell lines, xeroderma pigmentosum (XP) cell lines were found to be highly susceptible to homologous complement (C): cells were opsonized by C3 fragments on incubation with diluted normal human serum. C3 fragment deposition on XP cells was Ca2+-dependent and occurred on live cells but not UV-irradiated apoptotic cells. (Ca2+ is required for activation of the classical C pathway via C1q and the lactin pathway via mannose binding lectin (MBL), and the surface of apoptotic cells usually activates the alternative C pathway.) In this study we tested which of the pathways participates in XP cell C3 deposition. In seven cell lines that allowed C3 deposition (i), Clq was shown to be essential but MBL played no role in C activation, (ii) Cls but not MASP bound XP cells for activation, (iii) no antibodies recognizing XP cells were required for homologous C3 deposition, and (iv) the alternative pathway barely participated in C3 deposition. Furthermore, the levels of C-regulatory proteins for host cell protection against C, decay-accelerating factor (DAF, CD55) and membrane cofactor protein (MCP, CD46), were found to be relatively low in almost all XP cell lines compared with normal cells. These results indicate that XP cells activate the classical C pathway in an antibody-independent manner through the expression of a molecule which directly attracts C1q in a C-activating form, and that relatively low levels of DAF and MCP on XP cells facilitate effective C3 deposition. The possible relationship between the pathogenesis of XP and our findings is discussed.
Depletion of liver and splenic macrophages reduces the lethality of Shiga toxin-2 in a mouse model
Authors: PALERMO M S , ALVES ROSA M F , VAN ROOIJEN N , ISTURIZ M A
Published: March 2003

The haemolytic uraemic syndrome (HUS) is a clinical syndrome consisting of haemolytic anaemia, thrombocytopenia, and acute renal insufficiency. HUS is the most frequent cause of acute renal failure in childhood. It has been previously suggested that the presence of Shiga toxin (Stx) is necessary but not sufficient for HUS development, and cytokines such as tumour necrosis factor-alpha (TNF-α) and IL-1β appear to be necessary to develop the syndrome. Since the mononuclear phagocytic system (MPS) is the major source of these cytokines, macrophages might be one of the relevant targets for Stx action in the pathophysiology of HUS. In this study our objective was to examine the role of the hepatic and splenic macrophages in a mouse model of HUS induced by injection of Shiga toxin type-2 (Stx2) or Stx2 plus lipopolysaccharide (LPS). For this purpose, depletion of mice macrophages by liposome-encapsulated clodronate (lip-clod), followed by injection of STx2 or Stx2 plus LPS, was assayed. In this study we show that depletion of hepatic and splenic macrophages by clodronate treatment induces a survival of 50% in animals treated with Stx2 alone or in presence of LPS. This maximal effect was observed when lip-clod was injected 48–72 h before Stx2 injection. Biochemical and histological parameters show characteristics of the lesion produced by Stx2, discarding non-specific damage due to LPS or lip-clod. In addition, we determined that the toxic action of Stx2 is similar in BALB/c and N:NIH nude mice, indicating the T cell compartment is not involved in the Stx2 toxicity. Briefly, we demonstrate that macrophages play a central role in the pathophysiology of HUS, and that the systemic production of cytokines by liver and/or spleen is for Stx2 to manifest its full cytotoxic effect. In addition, the toxicity of Stx2 alone, or in presence of LPS, is independent of the T cell compartment.
Soluble CD40 in the serum of healthy donors, patients with chronic renal failure, haemodialysis and chronic ambulatory peritoneal dialysis (CAPD) patients
Authors: SCHWABE R F , ENGELMANN H , HESS S , FRICKE H
Published: March 2003

CD40 and its ligand CD40L are key players in T cell–B cell interaction and T cell–antigen-presenting cell (APC) interaction. Inhibition of CD40–CD40L interaction leads to severe humoral and cellular immunodeficiency. In this study we examined the presence of soluble CD40 (sCD40) in the serum of haemodialysis (HD) patients, CAPD patients, chronic renal failure (CRF) patients and healthy donors in order to evaluate the possible involvement of CD40 in uraemic immunodeficiency. Soluble CD40 was detected in the serum of healthy donors (n = 41) with a mean of 0.14 ± 0.12 ng/ml and in the urine of healthy donors with a mean of 1.80 ± 0.74 ng/ml. Soluble CD40 was highly elevated in all patients with impaired renal function. HD patients (n = 22) had up to 100-fold elevated sCD40 levels with a mean concentration of 8.32 ± 4.11 ng/ml, whereas CAPD patients (n = 10) had considerably lower levels of sCD40 with a mean of 3.58 ± 2.40 ng/ml. A strong correlation between sCD40 and serum creatinine levels was noted in CRF patients (n = 66). The highly elevated levels of sCD40 may point to the involvement of CD40 and its ligand CD40L in the clinical manifestation of uraemic immunodeficiency.
Increased activity of caspase 3 and caspase 8 in anti-Fas-induced apoptosis in lymphocytes from ageing humans
Authors: Aggarwal S , Gupta S
Published: March 2003

We have previously shown an increased susceptibility of T cell subsets to anti-Fas-induced apoptosis in human ageing [1]. In this study, we have examined the role of downstream mediators, including caspases, in Fas-mediated apoptosis in lymphocytes from ageing humans. The cleavage activity of caspase-8 and caspase-3 was compared between ageing and young subjects at different times following anti-Fas treatment, using colorimetric detection analysis. The expression of Fas-associated death domain (FADD), caspase-8, and caspase-3 in lymphocytes was compared at the protein level using Western blotting, and at the mRNA level by Northern blot analysis. In lymphocytes from ageing subjects, there was an early increase in the cleavage activity of caspase-8 and caspase-3 compared with young controls. Furthermore, increased protein expression of FADD, caspase-8 and caspase-3 at the basal level was observed in lymphocytes from ageing humans. Our results suggest that the altered expression and activity of molecules in the Fas/FasL signalling pathway may play a role in increased Fas-induced apoptosis and T cell deficiency in ageing humans.
Rheumatoid factors in primary Sjögren's syndrome (pSS) use diverse VH region genes, the majority of which show no evidence of somatic hypermutation
Authors: Elagib K E E , Børretzen M , Jonsson R , Haga H J , Thoen J , Thompson K M , Natvig J B
Published: March 2003

Rheumatoid factor (RF) is the most common autoantibody found in patients with Sjögren's syndrome (SS). To study the genetic origin and the mechanisms acting behind its generation we have characterized and sequenced the immunoglobulin VH genes used by 10 IgM RF MoAbs derived from peripheral blood of six female patients with pSS. We compared the structure of the RF immunoglobulin VH genes with those obtained previously from rheumatoid arthritis (RA) patients and healthy immunized donors (HID). VH1 and VH4 were each used by four RF clones, one clone was encoded by VH3 family gene and one by VH2 family gene. This distribution frequency was different from that observed in RA, where VH3 was the dominant family, followed by VH1. Eight different germ-line (GL) genes encoded the clones and all of these genes were seen previously in RA and/or HID RF. Five clones rearranged to JH6, four rearranged to JH4 and one to JH5, in contrast to RF from RA and HID, where JH4 was most frequently used. D segment use and CDR3 structure were diverse. Interestingly, three out of four VH4 clones used the GL gene DP-79 that was seen frequently in RA RF. The degree of somatic mutation in the pSS RF was very much lower than seen in RA and HID RF. All the pSS RF clones except three were in or very close to GL configuration. This indicates that there is little role for somatic hypermutation and a germinal centre reaction in the generation of RF from peripheral blood in pSS.
Antibody production in autoimmune BXSB mice. I. CD40L-expressing B cells need fewer signals for polyclonal antibody synthesis
Authors: Blossom S , Gilbert K M
Published: March 2003

Male, but not female, BXSB mice develop severe lupus associated with multiple immune system defects. It was recently shown that one immunological abnormality found in male BXSB mice encompasses B cell expression of CD40 ligand (CD40L) by an expanded population of large B cells. The present study was undertaken to determine how the CD40L-expressing large B cells in male BXSB mice compared with size-matched B cells from female mice in terms of their ability to secrete antibody. It was shown that the large B cells from female mice, similar to the small B cells from either male or female mice, required CD40 signalling, immunoglobulin cross-linking and cytokines for optimal antibody synthesis. In contrast, large B cells from male BXSB mice produced high levels of antibody when stimulated with only two of the three signals, and made significantly more total IgM and IgG, and anti-ssDNA antibody than size-matched B cells from female mice when stimulated with IL-4/IL-5 alone, IL-4/IL-5 plus low levels of anti-IgD-dextran, or IL-4/IL-5 plus anti-CD40 MoAb. The ability of the large B cells from male mice to produce antibody under suboptimal stimulatory conditions correlated with their expression of CD40L, and was inhibited by CD40-immunoglobulin. Taken together, these findings suggested that large CD40L-expressing B cells from male BXSB mice may be able to bypass a need for CD40 signalling from T cells, thus contributing to autoimmune disease by promoting antibody production in the absence of cognate T cell help.
The activation of the neutrophil respiratory burst by anti-neutrophil cytoplasm autoantibody (ANCA) from patients with systemic vasculitis requires tyrosine kinases and protein kinase C activation
Authors: Radford D J , Lord J M , Savage C O S
Published: March 2003

The ability of antineutrophil cytoplasm autoantibodies (ANCA) from patients with systemic vasculitis to stimulate protein kinase C (PKC) and tyrosine kinases was examined in human neutrophils. Using the superoxide dismutase-inhibitable reduction of ferricytochrome C, the kinetics of ANCA-induced superoxide (O2−) production were characterized and subsequently manipulated by specific inhibitors of PKC and tyrosine kinases. With this approach, ANCA IgG, but not normal IgG or ANCA F(ab′)2 fragments caused a time and dose dependent release of O2− from TNF-α primed neutrophils. The kinetics of ANCA-induced O2− production showed an initial 10–15 min lag phase compared to the N-formyl-l-methionyl-l-leucyl-l-phenylalanine response, suggesting differences in the signalling pathways recruited by these two stimuli. Inhibitor studies revealed that ANCA-activation involved members of both the Ca2+-dependent and -independent PKC isoforms and also tyrosine kinases. ANCA IgG resulted in the translocation of the βII isoform of PKC at a time corresponding to the end of the lag phase of O2− production, suggesting that PKC activity may be instrumental in processes regulating the activity of the NADPH oxidase in response to ANCA. Tyrosine phosphorylation of numerous proteins also peaked 10–15 min after stimulation with ANCA but not normal IgG. These data suggest that PKC and tyrosine kinases regulate O2− production from neutrophils stimulated with autoantibodies from patients with systemic vasculitis.
Benznidazole, a drug employed in the treatment of Chagas’ disease, down-regulates the synthesis of nitrite and cytokines by murine stimulated macrophages
Authors: Revelli S , LE. Page C L E , Piaggio E , Wietzerbin J , Bottasso O
Published: March 2003

Benznidazole (BZL) is a nitroheterocyclic drug employed in the chemotherapy of Chagas’ disease, a protozoan disease caused by Trypanosoma cruzi. Because this parasite mostly replicates in macrophages, we investigated whether BZL was likely to modify the synthesis of macrophage mediators such as nitrite, tumour necrosis factor-alpha (TNF-α), IL-1β, IL-6 and IL-10. Control and stimulated murine macrophages (lipopolysaccharide (LPS) and/or interferon-gamma (IFN-γ)) were treated with BZL and measurements were carried out in culture supernatants collected 24 h later. Synthesis of nitrite, IL-6 and IL-10 was maximal upon combined stimulation with LPS + IFN-γ, whereas lower amounts of the three mediators were detected when both stimuli were given alone. BZL treatment significantly reduced nitrite, IL-6 and IL-10 production, to undetectable levels in some cases, particularly IL-6 and IL-10. LPS was the most potent stimulus of IL-1β and TNF-α production, followed by LPS + IFN-γ and IFN-γ in decreasing order. BZL partly inhibited TNF-α synthesis, but this effect was smaller than that observed for nitrite, IL-6 and IL-10. LPS-induced production of IL-1β was also affected by BZL. Semiquantification of gene expression for inducible nitric oxide synthase (iNOS) showed that BZL completely inhibited iNOS gene induction by IFN-γ, and resulted in respective inhibitions of 30% and 50% with LPS- and LPS + IFN-γ-stimulated cells. BZL was not cytotoxic on macrophage cultures, as shown by the lactate dehydrogenase activity. Besides its trypanocidal activity, BZL may also alter the balance between pro- and anti-inflammatory mediators with important consequences for the course of T. cruzi infection.
Induction of keratinocyte IL-8 expression and secretion by IgG autoantibodies as a novel mechanism of epidermal neutrophil recruitment in a pemphigus variant
Authors: O’Toole E A , Mak L L , Guitart J , Woodley D T , Hashimoto T , Amagai M , Chan L S
Published: March 2003

A subset of pemphigus herpetiformis, a rare pemphigus variant, is characterized histopathologically by subcorneal acantholysis and neutrophilic infiltration. The mechanism of neutrophil infiltration is unknown, but chemokines such as IL-8 may play a role. We investigated the possible role of IL-8 in two such cases. Direct and indirect immunofluorescence studies demonstrated in vivo-bound and circulating IgG epithelial cell surface-binding autoantibodies, both predominated by IgG4 subclass. ELISA and immunoblotting studies revealed that the patients’ IgG autoantibodies recognized recombinant desmoglein 1 but not desmoglein 3. Preadsorption of the patients’ sera with recombinant desmoglein 1 completely removed the epidermal cell surface immunostaining. Significantly, immunohistochemistry demonstrated intense expression of IL-8, co-localized with in vivo-bound IgG, in the upper epidermis, where the acantholysis took place. Affinity-purified sera IgG from these two patients, a normal individual, and a pemphigus vulgaris patient containing desmoglein 1 autoantibodies, were incubated with normal human keratinocytes in vitro. Cells treated with these patients’ IgG secreted a seven-to-nine-fold increase of IL-8 (30–37 pg/ml) compared with the controls (2–4 pg/ml) and expressed a higher intensity of cytoplasmic IL-8 staining. These data demonstrate a novel functional role for IL-8 in the pathogenesis of the neutrophil-dominant subset of pemphigus herpetiformis. The autoantibody-induced epidermal cell IL-8 expression may represent a novel mechanism of epidermal neutrophil recruitment.
Induction of microthrombotic thrombocytopenia in normal mice by transferring a platelet-reactive, monoclonal anti-gp70 autoantibody established from MRL/lprmice: an autoimmune model of thrombotic thrombocytopenic purpura
Authors: Hashimoto K , Tabata N , Fujisawa R , Matsumura H , Miyazawa M
Published: March 2003

MRL/MpJ-lpr/lpr (MRL/lpr) mice spontaneously develop immune complex-mediated glomerulonephritis and thrombocytopenia. Although the presence of cross-reactive anti-phospholipid antibodies in sera of MRL/lpr mice has been demonstrated, possible relationships between detected autoantibodies and the development of thrombocytopenia have not been elucidated. Recent genetic analyses in a few different strains of lupus-prone mice have pointed out a close correlation between autoantibodies reactive with endogenous retroviral env gene product, gp70, and the development and severity of glomerulonephritis. In the process of establishing possibly nephritogenic anti-gp70 autoantibody-producing hybridoma cells from MRL/lpr mice, we identified an IgG2a-producing anti-gp70 hybridoma clone that induced microvascular intraluminal platelet aggregation, thrombocytopenia, and amenia upon transplantation into syngeneic non-autoimmune mice. This and two other anti-gp70 antibodies bound onto the surface of mouse platelets, and purified IgG2a of the anti-gp70 autoantibody induced glomerular lesions with characteristics of thrombotic thrombocytopenic purpura when injected into non-autoimmune mice. The pathogenic anti-gp70 autoantibody specifically precipitated a platelet protein with an approximate relative molecular mass of 40 000.
Fish oil feeding enhances lymphocyte proliferation but impairs virus-specific T lymphocyte cytotoxicity in mice following challenge with influenza virus
Authors: Byleveld M , Pang G T , Clancy R L , Roberts D C K
Published: March 2003

The effect of a fish oil diet on virus-specific cytotoxicity and lymphocyte proliferation was investigated. Mice were fed fish oil (17 g fish oil and 3 g sunflower/100 g) or beef tallow (17 g tallow and 3 g sunflower/100 g) diets for 14 days before intranasal challenge with influenza virus. At day 5 after infection, lung virus-specific T lymphocyte, but not macrophage or natural killer (NK) cell, cytotoxicity was significantly lower in mice fed fish oil, while bronchial lymph node cell proliferation to virus was significantly higher. In mice fed fish oil, spleen cell proliferation to virus was also significantly higher following immunization. The results showed that, despite improved lymphocyte proliferation, fish oil impairs primary virus-specific T lymphocyte cytotoxicity. This impairment may explain the delayed virus clearance that we have previously reported in infected mice fed the fish oil diet.
PERB11 (MIC): a polymorphic MHC gene is expressed in skin and single nucleotide polymorphisms are associated with psoriasis
Authors: Tay G K , Hui J , Gaudieri S , Schmitt-Egenolf M , Martinez O P , Leelayuwat C , Williamson J F , Eiermann T H , Dawkins R L
Published: March 2003

The susceptibility genes for psoriasis remain to be identified. At least one of these must be in the major histocompatibility complex (MHC) to explain associations with alleles at human leucocyte antigen (HLA)-A, -B, -C, -DR, -DQ and C4. In fact, most of these alleles are components of just two ancestral haplotypes (AHs) designated 13.1 and 57.1. Although relevant MHC gene(s) could be within a region of at least 4 Mb, most studies have favoured the area near HLA-B and -C. This region contains a large number of non-HLA genes, many of which are duplicated and polymorphic. Members of one such gene family, PERB11.1 and PERB11.2, are expressed in the skin and are encoded in the region between tumour necrosis factor and HLA-B. To investigate the relationship of PERB11.1 alleles to psoriasis, sequence based typing was performed on 97 patients classified according to age of onset and family history. The frequency of the PERB11.1*06 allele is 44% in type I psoriasis but only 7% in controls (Pc = 0.003 by Fisher's exact test, two-tailed). The major determinant of this association is a single nucleotide polymorphism (SNP) within intron 4. In normal and affected skin, expression of PERB11 is mainly in the basal layer of the epidermis including ducts and follicles. PERB11 is also present in the upper keratin layers but there is relative deficiency in the intermediate layers. These findings suggest a possible role for PERB11 and other MHC genes in the pathogenesis of psoriasis.
Oral administration of antigens from intestinal flora anaerobic bacteria reduces the severity of experimental acute colitis in BALB/c mice
Authors: Verdù E F , Bercik P , Cukrowska B , Farre-castany M A , Bouzourene H , Saraga E , Blum A L , Corthésy-Theulaz I , Tlaskalova-Hogenova H , Michetti P
Published: March 2003

Homeostasis between indigenous intestinal flora and host response may be broken in inflammatory bowel disease. The present study explores whether repeated oral administration of intestinal flora antigens can protect mice against dextran sodium sulphate (DSS)-induced colitis. Sonicates of Gram-positive, Gram-negative, or anaerobic resident bacteria isolated from mouse intestinal flora were fed to BALB/c mice by gastric gavage, with or without cholera toxin. After four weekly doses of 1 mg of these antigen preparations (or of PBS as control), DSS colitis was induced. One week later colitis was evaluated by clinical scores and histology. Mice fed a pool of the three sonicates had decreased inflammation scores (5 (1–14); median (range)) compared with PBS-fed control animals (15 (7–19); P < 0·05). Decreased inflammation was observed in mice fed anaerobic bacteria antigens (7 (6–11); P < 0·05 versus control), but not in mice fed a pool of Gram-positive and -negative sonicates (16 (12–16)). Inflammation scores of mice fed antigens with cholera toxin were similar to those of PBS-fed control animals. DSS-induced colitis can be suppressed by oral administration of normal intestinal flora antigens containing anaerobes.
Staphylococcal acid phosphatase binds to endothelial cells via charge interaction; a pathogenic role in Wegener’s granulomatosis?
Authors: Brons R H , Bakker H I , Van Wijk R T , Van Dijk N W , Muller Kobold A C , Limburg P C , Manson W L , Kallenberg C G M , Cohen Tervaert J W
Published: March 2003

The majority of patients with Wegener’s granulomatosis (WG) are chronic nasal carriers of Staphylococcus aureus. Chronic nasal carriage of S. aureus is associated with an increased risk of developing a relapse of the disease. The mechanism by which this occurs is still unknown. We hypothesized that a cationic protein of S. aureus, staphylococcal acid phosphatase (SAcP), acts as a planted antigen and initiates glomerulonephritis and vasculitis in patients with WG. In order to test the hypothesis that SAcP can act as a planted antigen in WG, we studied the ability of SAcP to bind to human umbilical vein endothelial cells (HUVEC) and human glomerular endothelial cells. We also studied whether this binding can be prevented by preincubation with an anionic protein, and whether binding of SAcP activates endothelial cells. We also evaluated whether antibodies in sera of patients with WG are able to bind to endothelial cell-bound SAcP. The results show that SAcP can act as a planted antigen by binding to both types of endothelial cells in a concentration-dependent manner. Binding of concentrations as low as 4 μg/ml can be detected on HUVEC within 5 min of incubation. Binding of SAcP to endothelial cells was charge-dependent but did not activate endothelial cells. Finally, endothelial cell-bound SAcP was recognized by sera of patients with WG. The data suggest a possible pathogenic role for SAcP by acting as a planted antigen thereby initiating glomerulonephritis and vasculitis in patients with WG.
Impact of highly active anti-retroviral therapy (HAART) on cytokine production and monocyte subsets in HIV-infected patients
Authors: Amirayan-Chevillard N , Tissot-dupont H , Capo C , Brunet C , Dignat-George F , Obadia Y , Gallais H , Mege J-L
Published: March 2003

HIV infection is associated with cytokine production by monocytes and expansion of a monocyte subset that expresses high levels of CD16. Our study was designed to investigate the effects of anti-retroviral therapies on these immune parameters. Four groups of HIV+ patients were included in the study. The first group comprised drug-naive patients (n = 20); the second included patients who received two inhibitors of HIV reverse transcriptase (n = 45); the third group received a therapy combining these two inhibitors and one inhibitor of HIV protease (HAART) (n = 35); the fourth consisted of patients who had stopped their treatment (n = 20). The release of inflammatory cytokines (tumour necrosis factor, IL-1β, IL-6) and immunoregulatory cytokines such as IL-10 by monocytes was determined by ELISA. The monocyte subsets expressing low or high levels of CD16 were studied by flow cytometry. Monocytes from patients naive of treatment released higher amounts of inflammatory cytokines and IL-10 than HIV− individuals. Each anti-retroviral therapy restored a normal pattern of cytokine secretion. Nevertheless, the release of cytokines increased again after the arrest of the treatment. The expansion of the monocyte subset that expresses high levels of CD16 was significantly decreased by HAART but not by the treatment including two inhibitors of reverse transcriptase. These results suggest that only HAART controls monocyte activation in the treatment of HIV infection.
Kinase mutant Btk results in atypical X-linked agammaglobulinaemia phenotype
Authors: Gaspar H B , Ferrando M , Caragol I , Hernandez M , Bertran J M , De gracia X , Lester T , Kinnon C , Ashton E , Espanol T
Published: March 2003

X-linked agammaglobulinaemia (XLA) is a B cell humoral abnormality arising from mutations in the gene encoding Bruton’s tyrosine kinase (Btk). The phenotype of XLA can be variable, with some individuals having a less severe immunophenotype, although in most cases this cannot be correlated with the Btk mutation or expression of Btk protein. In this study we describe clinical and immunological heterogeneity within the same pedigree. Analysis of the genetic defect identified a missense mutation in the kinase domain of Btk which, unusually, preserved Btk protein expression but at reduced levels, and also considerably diminished autophosphorylation activity. Structural analysis of the effect of this mutation on the kinase domain suggests that this mutation is not an integral part of the ATP or substrate binding domains but may affect the interaction of the kinase domain with its own kinase domain and other substrates. Together, these data may provide an explanation for the variable XLA phenotype.
Elevated serum concentrations of soluble CD14 in HIV− and HIV+ patients with tuberculosis in Africa: prolonged elevation during anti-tuberculosis treatment
Authors: Lawn S D , Labeta M O , Arias M , Acheampong J W , Griffin G E
Published: March 2003

Data are limited regarding serum concentrations of soluble CD14 (sCD14), a marker of macrophage activation, in patients with active tuberculosis (TB) and during drug treatment. In this study, concentrations of sCD14 were measured in serum samples obtained from 105 African subjects who were categorized into one of four groups: persons with pulmonary TB alone (TB+HIV−, n = 30), pulmonary TB and HIV co-infection (TB+HIV+, n = 20), or HIV infection alone (TB−HIV+, n = 25), and healthy controls (TB−HIV−, n = 30). Mean total sCD14 was significantly increased in serum of patients with newly diagnosed pulmonary TB (mean = 6·6 g/ml, s.d. = 1·6 g/ml) compared with healthy controls (mean = 3·1 g/ml, s.d. = 0·6 g/ml; P < 0·0001), and this elevation comprised proportionate increases in the α (2·1-fold greater, P < 0·0001) and β (2·0-fold greater, P < 0·0001) forms of sCD14. Total sCD14 was also increased in serum of HIV-infected patients (mean = 4·1 g/ml, s.d. = 1·9 g/ml; P < 0·01), but the highest concentrations were observed in patients with pulmonary TB and HIV co-infection (mean = 8·7 g/ml, s.d. = 3·1 g/ml; P < 0·0001). Analysis of serum samples prospectively collected from TB+HIV−patients during the first 3 months of successful anti-TB treatment demonstrated steep reductions in mean concentrations of the acute-phase protein, C-reactive protein, and the soluble lymphocyte activation marker, sCD25. In contrast, levels of sCD14 increased during the first month of treatment and slowly declined thereafter. These data indicate that the serum concentration of sCD14 is not a sensitive index of response to anti-TB treatment and suggest that cellular activation resolves more slowly in the macrophage pool compared with the lymphocyte pool during anti-TB treatment.
Peptide T does not ameliorate experimental autoimmune encephalomyelitis (EAE) in Lewis rats
Authors: Sáez-Torres I , Espejo C , Pérez J J , Acarín N , Montalban X , Martínez-Cáceres E M
Published: March 2003

Peptide T has been shown to inhibit T cell activation and cytokine production and function. Moreover, it has been reported to be a safe treatment in humans. We have studied the ability of peptide T to prevent or ameliorate EAE in Lewis rats. Peptide T was administered subcutaneously at different doses and phases of the disease according to several treatment protocols, but we could not observe a consistent effect of peptide T ameliorating the disease. Lymph node cell proliferation and IL-4 and interferon-gamma production were also studied. We conclude that peptide T neither prevents nor ameliorates EAE in Lewis rats.
Differences in regulatory pathways identify subgroups of T cell-derived Th2 cytokines
Authors: Rafiq K , Bullens D M A , Kasran A , Lorré K , Ceuppens J L , Van gool S W
Published: March 2003

We analysed regulatory mechanisms involved in the production of Th2 cytokines by freshly isolated human T cells. We used an in vitro culture system in which the primary signal was provided by a cross-linking anti-CD3 MoAb presented on the Fc receptors of P815 cells. Both CD80 and CD86, expressed on transfected P815 cells, were able to provide efficient costimulation for the production of IL-4, IL-5 and IL-13. IL-2 was also highly important for induction of all three Th2 cytokines. However, differences between IL-4 on the one hand and IL-5 and IL-13 on the other hand were observed when sensitivity to cyclosporin A (CsA) was studied. CsA (an inhibitor of calcineurin phosphatase activity) strongly inhibited IL-4 production, but it did either not affect or even increased IL-5 and IL-13 production. In accordance with this, CD80 and phorbol myristate acetate (PMA) (without anti-CD3 or calcium ionophore) were sufficient to induce production of IL-5 and IL-13, but not of IL-4. The subgrouping of Th2 cytokines was further confirmed at another level on the basis of differences in cell sources: IL-4 was predominantly produced by CD4+ T cells, while IL-5 and IL-13 were produced by both CD4+ and CD8+ T cells. Thus, differences in cell sources and in the requirement of the calcium/calcineurin-signalling pathway allowed us to identify two subgroups (IL-4 and IL-5/IL-13) among human Th2-type T cell cytokines.