bluuebunny/crossref_metadata_2025 huggingface dataset:
Chronic mucocutaneous candidiasis. I. Altered antigen-stimulated IL-2, IL-4, IL-6 and interferon-gamma (IFN-γ) production
Authors: Lilic D , Cant A J , Abinun M , Calvert J E , Spickett G P
Published: November 2003

Patients with chronic mucocutaneous candidiasis (CMC) present with persistent infections with the opportunistic yeast Candida. Impaired cell-mediated responses to Candida have been documented in CMC patients, but the defect remains poorly understood. The importance of Th1 cytokines in resistance and Th2 in susceptibility to Candida infections has recently been demonstrated in murine models. In our studies we evaluated production of IL-2 and IFN-γ (markers of Th1 type responses) as well as IL-4 and IL-6 (Th2 type markers) following stimulation with two kinds of Candida antigens (CAgs), polysaccharide antigens, tetanus toxoid and pokeweed mitogen. Our results demonstrate that CMC patients have impaired cytokine production upon in vitro stimulation with CAgs resulting in low or absent IL-2, increased IL-6 and either absent or increased IFN-γ production. Cytokine production following stimulation by other antigens was unaltered. The overall cytokine-producing capacity assessed through mitogen stimulation was also intact. Addition of IFN-α or IFN-γ to culture in an attempt to modify cytokine production did not have significant effects. Levels of soluble IL-6 receptors were not increased and could not account for increased IL-6 production. Our studies support the hypothesis that Candida antigens trigger a predominantly Th2 instead of a Th1 cytokine response in patients with CMC.
Chronic mucocutaneous candidiasis. II. Class and subclass of specific antibody responses in vivo and in vitro
Authors: Lilic D , Calvert J E , Cant A J , Abinun M , Spickett G P
Published: November 2003

Patients with chronic mucocutaneous candidiasis (CMC) succumb to persistent infections with the opportunistic yeast Candida. Impaired cell-mediated responses to Candida have been repeatedly reported while antibody responses were mostly found to be normal. The underlying defect remains poorly understood. It has recently been shown that CMC patients are also susceptible to infections with encapsulated bacteria, and may have associated IgG2 and IgG4 deficiency. Our previous studies demonstrated altered cytokine production in CMC patients. As cytokines can influence production and isotype of specific antibody, in 10 patients with CMC we measured the levels and isotype distributions of serum antibodies to Candida antigens (CAg), pneumococcal polysaccharide (PPS) and tetanus toxoid (TT) antigens. Peripheral blood lymphocytes were also stimulated in culture and the antibodies made in vitro were measured. Our data demonstrated that in vivo, CMC patients had very high levels of IgG and IgA CAg-specific antibodies. CAg-specific and PPS-specific IgG1 was markedly higher than in controls. Children but not adults with CMC had significantly lower levels of IgG2-specific antibody to CAg and PPS compared with age-matched controls. Patients had significantly higher levels of IgG3-specific antibody to all three antigens tested. These findings were in accordance with increased total IgG and IgG3 levels seen in CMC patients. In vitro, CMC patients, particularly children, did not respond as frequently to antigen stimulation as did their healthy controls. The level of specific antibody produced was also lower to all antigens tested, as was the amount of total immunoglobulins following antigenic and particularly mitogenic stimulation. Addition of interferon-alpha (IFN-α) or IFN-γ to cultures had variable, sometimes marked, effects. Our results demonstrate that CMC patients manifest subtle alterations in specific antibody responses to CAg, PPS and TT, which are most pronounced in children. This may relate to altered cytokine production also seen in these patients.
Crohn’s disease (CD) patients suffering from peripheral arthritis or ankylosing spondylitis reveal restricted T cell receptor Vβ regions in different temporal phases of disease
Authors: Lügering N , Kucharzik T , Fisahn M , Domschke W , Stoll R
Published: November 2003

Little is known about the mechanisms triggering and controlling both the development and perpetuation of extraintestinal complications in CD. The aim of the present study was to test the hypothesis that the T cell immune response in CD patients with joint complications may be altered when compared with patients without extraintestinal manifestations. We used a semiquantitative polymerase chain reaction assay to analyse the T cell antigen receptor repertoire in peripheral blood T cells from eight CD patients suffering from peripheral arthritis and ankylosing spondylitis, 12 CD patients without extraintestinal manifestations, and from seven non-CD patients with ankylosing spondylitis showing typical changes on joint radiographs. Being concerned that different patterns may be seen in different phases of the inflammatory disease process, we have also taken care to analyse sequential samples at various time points of the disease. Expression of all 22 Vβ genes was found in each healthy control and in each CD patient without extraintestinal manifestations and showed no major variation over time. Southern hybridization analysis of amplified products revealed a highly restricted Vβ repertoire in all CD patients suffering from peripheral arthritis and ankylosing spondylitis. In contrast, non-CD patients with ankylosing spondylitis without signs or symptoms of gastrointestinal problems demonstrated the presence of the entire Vβ repertoire. Our longitudinal studies confirmed variable Vβ usage over time, as certain transcripts were found only in distinct temporal phases of disease. Our data are not directly suggestive of a common superantigen model of CD, but instead emphasize a specific decrease in signals throughout the T cell receptor Vβ repertoire in CD patients suffering from joint complications.
Reduction in serum levels of antimitochondrial (M2) antibodies following immunoglobulin therapy in severe combined immunodeficient (SCID) mice reconstituted with lymphocytes from patients with primary biliary cirrhosis (PBC)
Authors: Abedi M R , Hammarström L , Broomé U , Angelin B , Edvard Smith C I , Christensson B
Published: November 2003

The effect of gammaglobulin treatment on autoantibody production was investigated in SCID mice reconstituted with human peripheral blood mononuclear cells (PBMC) obtained from patients with PBC. All reconstituted mice displayed the presence of human antimitochondrial antibodies (αM2Ab) of both IgG and IgM types before treatment with human immunoglobulin. Two weeks after i.p. injection of 20 ×106 PBMC into SCID mice, i.p. treatment with various preparations of human immunoglobulin was initiated. In control animals treated with saline, serum levels of human αM2Ab of the IgG type increased with time, peaking around 4 weeks after reconstitution. In contrast, human IgG autoantibodies rapidly decreased in all animals treated with human IgG. Treatment with a human IgM preparation had no effect on serum levels of αM2Ab of the IgG type. The results may suggest that the pronounced reduction of specific IgG autoantibodies was due to an increased catabolism of human IgG, including the autoantibodies, in the gammaglobulin-treated mice. Although the production of human αM2Ab in reconstituted mice could be easily shown, PBC-specific liver lesions or bile duct destruction were not observed, irrespective of treatment protocol.
Characterization of altered calcium signalling in T lymphocytes from patients with rheumatoid arthritis (RA)
Authors: Carruthers D M , Naylor W G , Allen M E , Kitas G D , Bacon P A , Young S P
Published: November 2003

Abnormal function of peripheral blood T lymphocytes is characteristic of RA; diminished proliferation and secretion of cytokines following in vitro mitogen stimulation are observed. We have investigated the calcium flux initiating T cell activation in rheumatoid peripheral blood mononuclear cells (PBMC) to determine whether abnormalities in signalling are also present. We have found that both phytohaemagglutinin (PHA-P)- and anti-CD3-stimulated calcium fluxes were much reduced in the patients’ PBMC compared with controls, with a mean six-fold difference (P < 0.01) in rate of Ca2+ flux with PHA-P stimulation. When purified T cells were examined with PHA and CD3 stimulation, a reduction in the peak and plateau [Ca2+]i was observed in RA T cells, but the rate of rise of [Ca2+]i was only reduced in those cells stimulated with PHA. These results suggest that alterations in the initiating signal may underlie the functional T cell abnormalities associated with RA, and that there may be an additional extrinsic influence from non-T cells in the PBMC population.
An analysis of apoptosis in lymphoid organs and lupus disease in murine systemic lupus erythematosus (SLE)
Authors: Ravirajan C T , Sarraf C E , Anilkumar T V , Golding M C H , Alison M R , Isenberg D A
Published: November 2003

Apoptosis is a programmed cell death process that helps to regulate both T cell and B cell development. In this study, we have investigated the levels of apoptotic death in cells of the thymuses and spleens (white matter) of autoimmune MRL-lpr/lpr mice with progressive lymphadenopathy and SLE disease activity; we also examined the renal pathology in these animals. Fas is a cell surface receptor, which when activated initiates the sequence of events that lead to apoptosis. In MRL-lpr/lpr mice Fas is defective, so the competency for apoptosis may be reduced. In young animals of advancing age the thymuses enlarged until in 5-month-old females the average weight was three times that at 1 month, and spleen and kidney weights also increased in size disproportionately. At light microscope level apoptotic cells in tissue sections were counted using both routine eosin and haematoxylin staining (to identify them by their morphology) and in situ end-labelling of cells with DNA strand breaks; their presence was further confirmed by electron microscopy. As the mice aged, the numbers of apoptotic cells in thymic cortex, thymic medulla and spleen white pulp areas reduced significantly (P < 0.01–0.001), whereas in BALB/c normal controls they increased significantly (P < 0.05). These changes were coincident with the development of severe lupus, whose activity was assessed by measuring serum anti-ssDNA and anti-dsDNA antibody titres and urinary protein (albumin) level which were elevated significantly by 5 months of age (P < 0.001 for both ssDNA and dsDNA and P < 0.01 for urine albumin) compared with their younger counterparts. Thus, lymphoid organ enlargement, decrease in apoptotic indices, elevated serum anti-ssDNA and anti-dsDNA antibody levels, and impaired renal function coincided with the onset and severity of lupus disease in lpr mice. It seems likely that there is a causal relationship between defective deletion of autoreactive lymphoid cells, imperfect Fas-mediated apoptosis and development of murine SLE.
Measurement of anticardiolipin antibodies by ELISA using β2-glycoprotein I (β2-GPI) in systemic sclerosis
Authors: Ihn H , Sato S , Fujimoto M , Kikuchi K , Igarashi A , Soma Y , Tamaki K , Takehara K
Published: November 2003

In order to determine the prevalence and clinical significance of β2-GPI-dependent anticardiolipin antibodies (β2-GPI/aCL) in patients with systemic sclerosis (SSc), serum samples from 80 patients with SSc, 20 patients with systemic lupus erythematosus (SLE), and 120 healthy control subjects were examined by ELISA using purified β2-GPI. IgG isotype β2-GPI/aCL was present in eight of 80 patients with SSc (10%), and the presence of β2-GPI/aCL IgG was significantly correlated with the presence of isolated pulmonary hypertension (PH). Furthermore, levels of β2-GPI/aCL IgG were significantly correlated with levels of mean pulmonary arterial pressure. These data suggest that IgG isotype β2-GPI/aCL might be a serological indicator of the severity of PH in patients with SSc.
CR1 stump peptide and terminal complement complexes are found in the glomeruli of lupus nephritis patients
Authors: Teixeira J E , Costa R S , Lachmann P J , Würzner R , Barbosa J E
Published: November 2003

The number of CR1 on podocytes is reduced in nephropathies with severe glomerular damage, especially in the diffuse proliferative glomerulonephritis (DPGN) of systemic lupus erythematosus (SLE). Reduction of CR1 number on erythrocytes is due to proteolysis of CR1 by macrophage proteases activated by the reaction of their complement receptors, which leaves a ‘CR1 stump peptide’ on the erythrocyte [1]. In the present study, we demonstrated the presence of the terminal complement complex (TCC) and the CR1 stump in histological sections of biopsies from patients with SLE by the indirect immunoperoxidase technique. Less severe glomerular lesions presented TCC deposits mainly in the mesangium (mesangial pattern). In lupus nephritis, with more severe glomerular damage, TCC deposits were detected both in the mesangium and in the capillary loops with podocyte involvement (mixed pattern). Patients with highly active DPGN presented a marked reduction of intact podocyte CR1 receptors in association with increased reactivity to the anti-CR1 stump antibody and with glomerular TCC deposits of mixed histological pattern. These results suggest that the decrease in the number of podocyte CR1 receptors in severe glomerular lesions of SLE may be due to a local proteolytic activity associated with activation and deposition of TCC.
Interferon-gamma (IFN-γ) can counteract the in vitro inhibitory effect of an HIV p24 immunosuppressive heptapeptide
Authors: Luzzati A L , Boirivant M , Giacomini E , Giordani L , Di Modugno F , Chersi A
Published: November 2003

Previous work from our laboratory demonstrated that a synthetic heptapeptide (Ch7), corresponding to a conserved sequence of HIV core protein p24 (aa 232–238), was able to specifically abrogate antigen-induced responses in cultures of normal human peripheral blood lymphocytes (PBL). In the present study we show that Ch7 did not inhibit the induction of IFN-γ-secreting cells nor the accumulation of IFN-γ mRNA in antigen-stimulated cultures. However, delayed addition of recombinant human IFN-γ to Ch7-suppressed cultures was able to restore fully the capacity to mount an antigen-specific antibody response. Thus, although the Ch7 immunosuppressive effect may not be directly related to a decreased production of IFN-γ, an increased level of this cytokine is certainly able to counteract the negative effect of the peptide.
Malaria, blood glucose, and the role of tumour necrosis factor (TNF) in mice
Authors: Elased K M , Taverne J , Playfair J H L
Published: November 2003

Hypoglycaemia in falciparum malaria is associated with a poor prognosis and is correlated with mortality. High levels of serum TNF are also correlated with disease severity and mortality, and it has been suggested that TNF may cause the hypoglycaemia. However hypoglycaemia in mice infected with Plasmodium chabaudi or the lethal strain of P. yoelii YM is related to hyperinsulinaemia. Its development was not prevented by treatments which diminished TNF activity or production without affecting levels of plasma insulin. Conversely, it was inhibited by diazoxide, which inhibited insulin secretion but did not affect TNF production. Furthermore, in mice exhibiting neurological symptoms during infection with P. berghei, blood glucose concentrations were significantly raised when TNF levels were high, and TNF levels in the spleen were highest of all in non-lethal P. yoelii infections in which hypoglycaemia does not occur. Administration of human rTNF to normal animals caused an increase rather than a drop in blood glucose levels. Mice transgenic for human TNF did not develop hypoglycaemia when infected with P. yoelii YM, but showed signs of insulin resistance. In line with current views on the role of TNF in obesity and the control of glucose homeostasis, we conclude that the hypoglycaemia of malaria is not caused by increased levels of TNF, which may in fact be beneficial, but is secondary to a hyperinsulinaemia that is probably stimulated directly by products of the parasite.
Expression of Haemophilus influenzae type b idiotype 1 on naturally acquired antibodies
Authors: Ulanova M , Hahn-Zoric M , Lau Y L , Lucas A , Hanson L Å
Published: November 2003

The Chinese population in Hong Kong has a low incidence of invasive Haemophilus influenzae type b (Hib) disease, as well as carriage of the microorganism. Likely stimuli for the natural antibodies to Hib, which might protect against Hib infection, are cross-reactive antigens of bacteria like Escherichia coli K100. Our aim was to determine the isotype and idiotype distribution and cross-reactivity of natural antibodies against Hib capsular polysaccharide (CP) in healthy Hong Kong Chinese. Titration of 20 sera by ELISA showed IgG antibodies reacting with Hib CP in all individuals. The antibodies were mainly IgG2, and their avidity index ranged widely. Isoelectric focusing (IEF) combined with immunoblotting showed patterns of IgG2 antibody clones against the CP of Hib and E. coli K100 which were similar in 10 cases. Absorption with Hib CP only eliminated some bands in two sera. Absorption with K100 CP did not remove any anti-Hib CP bands. In three sera additional clones of antibodies reacting to K100 CP only, disappeared after absorption with this CP. Spectrotypic analyses of IgG antibodies reacting with anti-Hib idiotype 1 (Id-1) revealed stronger IEF patterns with bands in differing locations compared with anti-Hib CP antibodies. The strong reactivity of serum IgG, IgA and IgM antibodies with monoclonal anti-Hib Id-1 was confirmed by ELISA. This reactivity was not abolished after absorption of the sera with either Hib CP, or K100 CP. The data indicate a high prevalence of Id-1 among Hong Kong Chinese. However, only one individual had Id-1 antibodies specific for Hib CP, judging from absorption experiments. Others had much lower activity of Id-1 anti-Hib CP antibodies compared with the total IgG Id-1, suggesting that Hong Kong subjects have Id-1-positive antibodies in their serum which are not specific for Hib CP. This is consistent with the nature of Id-1, which is a marker of A2VL region usage rather than a marker of a Hib CP paratope. We suggest that natural antibodies reacting with Hib CP in healthy Hong Kong Chinese are the product of exposure to some cross-reactive antigen(s), different from both Hib and E. coli K100 CP.
T cell responses to a mixture of Mycobacterium tuberculosis peptides with complementary HLA-DR binding profiles
Authors: Jurcevic S , Hills A , Pasvol G , Davidson R N , Ivanyi J , Wilkinson R J
Published: November 2003

The T cell response to a mixture of eight peptides derived from sequences of the Mycobacterium tuberculosis 16-, 19- and 38-kD antigens (MTBmix-8) has been studied. The peptides were selected on the basis of complementary binding to nine HLA-DR molecules (HLA-DR1 to DR9). MTBmix-8 at 6.25 and 50 μg/ml gave rise to significant stimulation (P < 0.05) of peripheral blood mononuclear cells (PBMC) from healthy tuberculin-positive and both untreated and treated diseased subjects, but not in any of a control group of healthy tuberculin-negative subjects. MTB-mix-8 stimulated proliferation of PBMC from healthy tuberculin-positive individuals at lower concentrations than the individual component peptides. However, the maximal stimulation achieved was only slightly higher than that achieved with individual peptides. MTBmix-8 also stimulated the production of interferon-gamma (IFN-γ) in vitro. Using the mean ± 2 s.d. of the values for IFN-γ production in the tuberculin-negative population as a cut-off, MTBmix-8 at 6.25 μg/ml was able to detect infection with a sensitivity of 100% in untreated patients, 87% in treated patients, and 82% in tuberculin-positive controls. The corresponding figures for the most potent single peptide (16p91–110) were: 66% in untreated patients, 71% in treated patients and only 42% in controls. Thus, using the IFN-γ-based assay, which has the additional advantages of speed and does not require radioactivity, the mixture of peptides is more sensitive than single peptides in diagnosing infection.
T helper type-1 (Th1)/Th2 profiles of peripheral blood mononuclear cells (PBMC); responses to antigens of Chlamydia trachomatis in subjects with severe trachomatous scarring
Authors: Holland M J , Bailey R L , Conway D J , Culley F , Miranpuri G , Byrne G I , Whittle H C , Mabey D C W
Published: November 2003

Increased stimulation of Th2 cytokines may contribute to the development of persistent ocular chlamydial infection, resulting in the blinding pathological changes of trachoma. Proliferation and cytokine production profiles of PBMC in response to stimulation with antigens of Chlamydia trachomatis were compared in 30 patients with severe conjunctival scarring due to trachoma and in 30 age-, sex- and location-matched controls. Interferon-gamma (IFN-γ) and IL-4 were detected at the single-cell level by ELISPOT assay. Transcription of the genes encoding IFN-γ, IL-4 and IL-10 was detected in mRNA isolated from parallel cultures of PBMC using reverse transcriptase-polymerase chain reaction (RT-PCR). Incubation with the chlamydial heat shock protein (hsp)60 resulted in increased numbers of IL-4-producing cells in PBMC isolated from patients with scarring disease and increased secretion of IFN-γ from PBMC of control subjects. Incubation with the chlamydial major outer membrane protein (MOMP) increased the number of IFN-γ-producing cells in the control group only. Messenger RNA encoding IL-4 was only detected in PBMC of patients with scarring disease after in vitro stimulation with chlamydial antigens, but IFN-γ mRNA and IL-10 mRNA were also more frequently detected in this group. Thirty-eight subjects were HLA-DRB1 and -DQB1 typed. Associations were observed between certain HLA class II alleles and cellular immune responses to chlamydial antigens. No HLA associations were found with clinical status, and overall we found no evidence of strong associations and the type of immune response. These data are consistent with a role for Th2 cells and cytokines in the pathogenesis of trachomatous scarring.
Production of IL-5 and IL-6 by peripheral blood mononuclear cells (PBMC) from patients with Echinococcus granulosus infection
Authors: Riganò R , Profumo E , Teggi A , Siracusano A
Published: November 2003

The role of Th2 cytokines in human hydatidosis was evaluated in ELISA determining IL-5 and IL-6 production in PBMC cultures from 27 pharmacologically treated hydatid patients and from 13 uninfected controls. PBMC from patients produced large amounts of parasite antigen-driven IL-5, whereas PBMC from uninfected individuals produced none. In contrast, PBMC from patients and from uninfected controls produced large amounts of parasite antigen-driven IL-6. Immunoglobulin isotype analysis revealed that IL-5 production correlated significantly with IgE and IgG4 expression (IL-5/IgE r=0.5, P<0.05; IL-5/IgG4 r=0.6, P<0.05). The high IL-5 levels in supernatants from patients’ PBMC did not correspond to an increase in eosinophils. Neither IL-5 nor IL-6 production showed an association with the outcome of therapy. Overall, these findings confirm that the lymphocytes of individuals with Echinococcus granulosus infection contain Th2-like subpopulations.
Anti-RNA polymerase antibodies in systemic sclerosis (SSc): association with anti-topoisomerase I antibodies and identification of autoreactive subunits of RNA polymerase II
Authors: Harvey G R , Rands A L , McHugh N J
Published: November 2003

The prevalence of autoantibodies to the three RNA polymerase (RNAP) enzymes in the sera of 249 SSc patients was measured using the technique of immunoprecipitation of 35S-methionine-labelled K562 cell extracts. Forty-six anti-RNAP sera were detected (18.5%) and three main groups were identified: anti-RNAP I/III sera (10; 4.0%), anti-RNAP I/II/III sera (15; 6.0%), and sera precipitating the phosphorylated (IIO) form of RNAP II (18; 7.2%). All sera in the third group also precipitated topoisomerase I (topo I), and six of them also precipitated the unphosphorylated (IIA) form of RNAP II. Although RNAP II/topo I multienzyme complexes may occur in cell extracts, autoreactive epitopes were shown to be located on both enzymes by a combination of antigen depletion studies, and in vitro assays which demonstrated functional inhibition of topo I activity. Furthermore, immunoblotting experiments using affinity-purified extracts demonstrated that all sera with anti-RNAP II antibodies recognized the largest RNAP II subunit in its phosphorylated form (IIo; 240 kD), whereas the unphosphorylated subunit (IIa; 220 kD) was only recognized by sera which also precipitated RNAP IIA. Therefore at least two different sites on the largest subunit of RNAP II are recognized by SSc sera, and one of these sites is unique to the phosphorylated (IIO) form.
Lung granulomatous response induced by infection with the intestinal nematode Nippostrongylus
brasiliensis is suppressed in mast cell-deficient Ws/Ws rats
Authors: Arizono N , Nishida M , Uchikawa R , Yamada M , Matsuda S , Tegoshi T , Kitamura Y , Sasabe M
Published: November 2003

Certain nematode infections induce eosinophil infiltration and granulomatous responses in the lungs. To examine the role of mast cells in the development of lung lesions, normal +/+ and genetically mast cell-deficient Ws/Ws rats were infected with the nematode Nippostrongylusbrasiliensis. In +/+ rats, numbers of eosinophils in bronchoalveolar lavage fluid (BALF) increased significantly 3–7 days after infection, and granulomatous responses composed of histiocytes/macrophages and multinucleate giant cells were triggered in the lungs 3–14 days after infection. Challenge infection, which was carried out on day 28 after primary infection, induced much higher levels of granulomatous response than after primary infection, suggesting that the response is mediated at least in part by an immunological mechanism. In Ws/Ws rats, both the eosinophil percentage in BALF and the size of the granulomas in the lungs were significantly smaller than in +/+ rats after primary as well as after challenge infection. The amount of rat mast cell protease (RMCP) II in +/+ rat BALF was increased 1 day after primary infection and more significantly after challenge infection, suggesting that lung mucosal mast cells were activated more markedly after the challenge infection. In Ws/Ws rats, RMCP II was undetectable throughout the observation period. The time course of nematode migration in the lungs did not differ in +/+ and Ws/Ws rats. These results suggest that mast cell activation might be relevant to eosinophil infiltration and granulomatous response in the lungs, although the responses do not affect lung migration of the nematode.
Inhibition of natural killer (NK) cell activity against varicella-zoster virus (VZV)-infected fibroblasts and lymphocyte activation in response to VZV antigen by nitric oxide-releasing agents
Authors: Ito M , Watanabe M , Kamiya H , Sakurai M
Published: November 2003

The addition of nitric oxide (NO)-releasing agents, S-nitroso-N-acetyl-DL-penicillamine (SNAP), 1-hydroxy-2-oxo-2,3-bis(2-aminoethyl)-1-triazene (NOC18), 30{(±)-(E)-ethyl-2′-[(E)-hydroxyimino]-5-nitro-3-hexenecarbamoyl}-pyridine (NOR4) significantly inhibited NK cell activity against VZV-infected cells, while antibody-dependent cell-mediated cytotoxicity (ADCC) against VZV-infected cells was unaffected. Interferon-alpha (IFN-α) production by non-adherent peripheral blood mononuclear cells (NPBMC) cultured with VZV-infected cells was decreased by the addition of NO-releasing agents. Lymphocyte proliferation and the expression IL-2 receptor (CD25) in response to VZV antigen were also inhibited by the addition of NO-releasing agents. These results suggest that the production of NO by an inflammatory process may lead to inhibition of NK cell- and T cell-mediated immunity to VZV infection.
Characterization of type 1 and type 2 cytokine production profile in physiologic and pathologic human pregnancy
Authors: Marzi M , Vigano A , Trabattoni D , Villa M L , Salvaggio A , Clerici E , Clerici M
Published: November 2003

Antigen- and mitogen-stimulated cytokine production by peripheral blood mononuclear cells (PBMC) of 50 pregnant women and 31 age- and sex-matched non-pregnant controls were analysed to determine whether changes in cytokine production occur during normal and pathologic human gestation. The pregnant women, consecutively enrolled during a 3-month period, were undergoing a normal, non-pathologic pregnancy at the time of entry into the study, and underwent ultrasound examination to ascertain the exact week of pregnancy and the vitality of the fetus. Forty of the 50 pregnancies (80%) terminated physiologically with the birth of normal babies. Spontaneous abortions were observed in 5/50 (10%) women, and five women gave birth to newborns small for gestational age (SGA). A decrease in the production of IL-2 and interferon-gamma (IFN-γ) accompanied by an increase in production of IL-4 and IL-10, was observed in normal pregnancy, with the lowest quantities of IL-2 and IFN-γ and the highest quantities of IL-4 and IL-10 present in the third trimester of pregnancy. Statistically significant increased production of both IL-2 and IFN-γ and reduced production of IL-10 characterized pathologic pregnancies and distinguished them from normal pregnancies. These preliminary data suggest that a type 2 cytokine profile may be associated with normal human pregnancy, whereas the lack of a dominant type 2 cytokine profile may be indicative of a pathologic pregnancy.
Role of macrophages in the development of arteritis in MRL strains of mice with a deficit in Fas-mediated apoptosis
Authors: Taniguchi Y , Ito M R , Mori S , Yonehara S , Nose M
Published: November 2003

The lpr and gld genes have been shown to encode the Fas antigen deletion mutant and the Fas ligand (FasL) mutant, respectively. An MRL strain of mice bearing the gld gene was observed to spontaneously develop granulomatous arteritis, similar to that in mice bearing the lpr gene, indicating that arteritis in this strain is due to an inefficient Fas–FasL interaction resulting in an incapacity for Fas-mediated apoptosis. The arterial lesions in both strains were characterized by a remarkable perivascular accumulation of activated macrophages bearing Mac-2 antigen, following the infiltration of CD4+ cells, and this resulted in the destruction of the arterial wall. Almost all of these infiltrating cells were Fas-positive, as determined in MRL/gld mice. Macrophage colony-stimulating factor (M-CSF), which is present at increased levels in MRL/lpr mice, but not in MRL/Mp- +/+ (MRL/+) mice, induced the expression of Mac-2 antigen and Fas antigen on spleen adherent cells of MRL/+ mice. Moreover, continuous infusion of M-CSF into the peritoneal cavity or subcutis of MRL/+ mice induced the release of oxygen radicals of peritoneal macrophages or granuloma formation associated with the massive accumulation of Mac-2+ cells, respectively. These findings suggest that macrophages in these mice, which may be activated by M-CSF and may avoid Fas-mediated apoptosis, play a critical role as effector cells in the destruction of arterial wall.
Epitope analysis of the Goodpasture antigen using a resonant mirror biosensor
Authors: Levy J B , Turner A N , George A J T , Pusey C D
Published: November 2003

We have used a new technique for studying molecular interactions—a resonant mirror biosensor—to identify B cell epitopes within the Goodpasture antigen, which has recently been identified as the non-collagenous domain of the α3-chain of type IV collagen (α3(IV)NC1). Recombinant antigen (r-α3) was immobilized onto the sensing surface of a sample cuvette, and the binding of patients’ autoantibodies or a MoAb to the Goodpasture antigen was followed in real time. All patients’ sera bound r-α3 in this system, while control sera did not bind. A MoAb inhibited the binding of all patients’ autoantibodies to r-α3, from 27% to 90% (mean inhibition 60%), and patients’ sera cross-inhibited the binding of each other to the antigen. Binding was inhibited by pre-incubation of autoantibody with both native sheep α3(IV)NC1 and purified human α3(IV)NC1 monomers. Inhibition experiments using soluble overlapping peptides from human α3(IV)NC1 identified putative B cell epitopes. These results suggest that there is a major immunodominant epitope on the Goodpasture antigen, and that there is very limited heterogeneity in the autoantibody response in Goodpasture’s disease. The resonant mirror biosensor can be successfully used to monitor antibody–antigen binding using polyclonal sera, and to map epitopes on autoantigens.
Diminished interferon-gamma (IFN-γ) production by bacterial antigen-specific T cells in atopic patients
Authors: Shimojo N , Kohno Y , Katsuki T , Hoshioka A , Honma K , Saito K , Niimi H
Published: November 2003

In this study, we established and studied cytokine production of T cell lines (TCL) specific to either a purified protein derivative of Mycobacterium tuberculosis (PPD) or Dermatophagoides farinae (Df) from atopic patients and non-atopic healthy subjects. IFN-γ was detected in the culture supernatants of all of 36 PPD-specific TCL established from healthy controls, whereas only 24 of 38 PPD-specific TCL from patients produced IFN-γ. Furthermore, the amounts of IFN-γ produced by PPD-specific TCL from patients were significantly lower than those from healthy controls. No IL-4 was detected in any PPD-specific TCL from either healthy controls or atopic patients. The amounts of IL-4 production from Df-specific TCL from atopic patients were much higher than from healthy controls, while few TCL produced IFN-γ. These results suggest that the skewing to the Th2-type T cell response in atopic patients is a response not only to allergens, but also to bacterial antigens, compared with non-atopic subjects. Activation of PPD-specific TCL from patients with calcium ionophore A23187 plus phorbol myristate acetate resulted in much higher IFN-γ production than in TCL established from healthy controls, indicating that the low production of IFN-γ by PPD-specific T cells from atopic patients is not due to an intrinsic T cell defect but to some regulatory mechanisms.
The mode of action of treatment by IgG of haemolytic anaemia induced by an anti-erythrocyte monoclonal antibody
Authors: Pottier Y , Pierard I , Barclay A , Masson P L , Coutelier J-P
Published: November 2003

In order to gain insight into the mechanisms by which the infusion of IgG can improve some autoimmune diseases, we induced haemolytic anaemia in mice by the injection of anti-erythrocyte MoAbs derived from NZB mice by S. Izui (Geneva). The IgG1 antibody 31-9D induces anaemia by erythrocyte sequestration in the spleen and liver, whereas the IgG2a antibody 34-3C triggers erythrophagocytosis (Shibata et al., Int Immunol 1990; 2:1133). Treatment of mice with pools of either human or mouse IgG clearly attenuated the anaemia induced by 34-3C, but not by 31-9D. Similar protection was obtained with human monoclonal IgGs from myeloma patients. Prior absorption by mouse erythrocytes did not affect the efficacy of the injected IgG. Treatment with Fc fragments also reduced the anaemia. In vitro experiments confirmed that 34-3C, but not 31-9D, triggered erythrocyte phagocytosis by murine macrophages. This process was completely inhibited by addition of polyclonal or myeloma IgG or of human Fc fragments. These results indicate that, in this model of autoimmune pathology, the protective effect of IgG is mediated by its interaction with the macrophage Fc receptors.
IL-1 receptor antagonist in saliva; characterization in normal saliva and reduced concentration in Sjo¨gren's syndrome (SS)
Authors: Dubost J-J , Perrier S , Afane M , Viallard J-L , Roux-Lombard P , Baudet-Pommel M , Begue C , Kemeny J-L , Sauvezie B
Published: November 2003

The characterization of a salivary factor cross-reacting with IL-1 receptor antagonist (IL-1Ra) is described. The apparent molecular weights of two species were 23 kD, consistent with the secreted peptide (sIL-1Ra), and 20 kD, consistent with the intracellular peptide (icIL-1Ra). It had an inhibitory activity on IL-1-stimulated fibroblasts, which is characteristic of IL-1Ra. Its source was the oral mucosa and not the salivary glands. Saliva from patients with SS contained significantly less IL-1Ra than saliva from controls. The decrease was marked in patients with early dental loss but whose xerostomia was still partial. In SS, the salivary IL-1/IL-1Ra imbalance may promote inflammatory lesions in the mouth and impede mucosal cell differentiation.
Abundance of intraepithelial γδ T cells in hypertrophic obstructive but not in chronically infected adenoids
Authors: Olofsson K , Hellstroøm S , Hammarstroøm M-L
Published: November 2003

Using quantitative morphometric analysis of immunohistochemically stained tissue sections we compared hypertrophic obstructive adenoids (HOA, n = 10) from children without middle ear disease with chronically infected adenoids (CIA, n = 10) from children with middle ear disease. γδ T cell receptor (TCR)+ cells constituted the dominating T cell population in the surface epithelium of HOA, while αβ TCR+ cells were the dominating intraepithelial T cell population in CIA. Intraepithelially CD8+ cells dominated over CD4+ cells in both diseases. Intraepithelially B cells were not detected. The cellular composition of follicles, with B cells dominating followed by activated CD4+αβ TCR+ cells, was the same in both groups. However, the number of follicles in CIA was twice as many as in HOA. In the deeper interfollicular areas granulocytes were more abundant in CIA than in HOA. The latter two findings suggest a more pronounced inflammatory response in the adenoids of patients with middle ear disease. There was no significant difference with regard to pathogenic bacterial strains colonizing the adenoid surface when comparing the two patient groups. These results suggest that in patients with HOA γδ TCR+ T cells help to maintain the integrity of the surface epithelium, thereby preserving its protective function. On the basis of our results we speculate that CIA have a malfunctioning defence, thereby facilitating long-standing infections deep in the adenoid. This may be the main reason for development of middle ear disease and an indication for adenoidectomy in patients with CIA.
Reduced IL-10 secretion by CD4+ T lymphocytes expressing mutant cystic fibrosis transmembrane conductance regulator (CFTR)
Authors: Moss R B , Bocian R C , Hsu Y-P , Dong Y-J , Kemna M , Wei T , Gardner P
Published: November 2003

Expression of the CFTR protein is thought to be physiologically important only in exocrine epithelial cells. However, chronic respiratory inflammation and infection remain unexplained phenomena in disease pathogenesis. Non-transformed, antigen-responsive CD4+ T cells cloned from healthy controls and CF patients homozygous or heterozygous for the δF508 mutation transcribed CFTR mRNA and expressed immunoreactive cytoplasmic CFTR protein. T cell clones (TCC) from controls and CF patients displayed equivalent Ca2+-mediated Cl− current; however, TCC from patients with CF but not controls displayed defective cAMP-mediated Cl− current. Although CF-derived TCC preserved mitogen and antigen proliferative responses and specificity to tetanus toxoid epitopes, they selectively secreted ≈ 45% less IL-10 compared with control TCC after activation with concanavalin A (Con A) (624 ± 101 versus 1564 ± 401 pg/ml per 106 cells, respectively; P = 0.04) or anti-CD3/phorbol ester (5148 ± 1634 versus 11 788 ± 2390 pg/ml; P = 0.05). This difference was independent of atopy. Secretion of interferon-gamma, IL-2, and IL-4 was comparable in CF and control TCC after both forms of activation, while IL-5 was reduced in CF TCC following anti-CD3/phorbol myristate acetate (PMA) but not after Con A. We conclude that expression of mutant CFTR in human TCC is accompanied by ion channel dysfunction characteristic of the CF phenotype, and is accompanied by a reduction in IL-10 secretion after polyclonal activation. It is possible that disruption of IL-10-mediated anti-inflammatory homeostasis may contribute to early onset sustained inflammation in CF airways.
Diabetic sera react with the glutamic acid decarboxylase molecule in a dimeric-oligomeric form
Authors: Rowley M J , Chen Q Y , Teoh K L , Zimmet P Z , Tuomi T , Knowles W J , Mackay I R
Published: November 2003

Glutamic acid decarboxylase (GAD) is a major autoantigen in insulin-dependent diabetes mellitus (IDDM). This was initially identified as a 64–65 kD molecule according to migration in gels after immunoprecipitation from pancreatic islets. We studied the antigenicity of two different radiolabelled preparations of GAD, derived either by affinity purification from porcine brain and known to contain GAD 65 and GAD 67, or by expression from a cDNA for human GAD 65 by rabbit reticulocyte lysate (RRL). Radiolabelled immunoprecipitated pellets from the reaction of potent antisera to GAD from patients with IDDM were examined by autoradiography after SDS–PAGE under reducing or non-reducing conditions. Also, preparations of porcine brain GAD were ‘depleted’ of GAD by exposure to antisera, and then similarly re-examined. Autoradiography of radiolabelled GAD either affinity purified from porcine brain, or expressed by RRL, showed that the immunoprecipitated protein migrated under non-reducing conditions according to a Mr of ∼110–130 kD, corresponding to dimeric forms of monomeric GAD of ∼55–65 kD. Depletion by immunoprecipitation of this minor higher Mr component from preparations of GAD left, in the supernatant, an abundance of GAD of Mr 64–65 kD corresponding to monomer that was completely non-reactive with potent IDDM sera. We conclude that IDDM sera react with the GAD molecule in a dimeric (or oligomeric) form. Our findings have general connotations for self-tolerance and autoimmunity.
Primary localized amyloidosis of the eyelid: two cases of immunoglobulin light chain-derived proteins, subtype λV respectively λVI
Authors: Olsen K E , Sandgren O , Sletten K , Westermark P
Published: November 2003

Primary localized amyloidosis has been described in many different organs in the body. Studies by immunohistochemical techniques have suggested an immunoglobulin light chain origin of the amyloid material. Only in a limited number of cases has the amyloid protein been characterized by amino acid sequence analysis as subtypes of immunoglobulin light chain or heavy chain. In this report, two cases of primary localized amyloidosis of the eyelid are presented. The amyloid substance has been extracted and a major fibril protein subjected to amino acid sequence analysis. Both amyloid proteins were part of the variable region of immunoglobulin light chains, subtype λV and subtype λVI, respectively. While λVI has been shown to be a common subtype in systemic immunoglobulin light chain-amyloidosis, it has never been demonstrated in localized amyloid. Very few λV immunoglobulin light chains have been characterized and the subgroup has never been found in amyloid before.
IL-1β production by human polymorphonuclear leucocytes stimulated by anti-neutrophil cytoplasmic autoantibodies: relevance to systemic vasculitis
Authors: Brooks C J , King W J , Radford D J , Adu D , McGrath M , Savage C O S
Published: November 2003

Neutrophils accumulate in the acute blood vessel lesions of patients with autoimmune systemic vasculitis. They have been shown previously to produce the cytokine IL-1β in response to stimulation with TNF. This study demonstrates that neutrophils can be stimulated by anti-neutrophil cytoplasmic antibodies (ANCA), which are present in patients with systemic vasculitis, to express mRNA and protein for IL-1β. Both human ANCA and MoAbs to a variety of autoantigens recognized by ANCA, including proteinase 3, myeloperoxidase, bactericidal/permeability increasing protein and elastase, are effective. This response can be inhibited by actinomycin and cycloheximide, suggesting a requirement for de novo protein synthesis. IL-1β production can be inhibited by pooled human intravenous immunoglobulins but not by FK506 or cyclosporin A. These data suggest that ANCA in patients with active vasculitis may stimulate neutrophils to produce cytokines. It is hypothesized that cytokine production from neutrophils that accumulate in significant numbers in vasculitic lesions contribute to and augment the local inflammatory response by the activation of vascular endothelial cells and infiltrating leucocytes.
Structural peculiarities of a truncated VκIII immunoglobulin light chain in myeloma with light chain deposition disease
Authors: Decourt C , Cogné M , Rocca A
Published: November 2003

We report on the primary sequence of the monoclonal immunoglobulin light chain (LC) REV involved in myeloma-associated light chain deposition disease (LCDD). This sequence was deduced from that of the corresponding complementary (c)DNA in bone marrow plasma cells. Products of three independent amplifications by polymerase chain reaction (PCR) were sequenced and found to be identical. The κ mRNA encoding this N-glycosylated LC showed an overall normal structure consisting of a VκIII segment rearranged to JκII. Direct N-terminal amino acid sequencing of the circulating monoclonal IgA2,κ showed identity with the bone marrow-derived sequence. The κ-chain presented several unusual features affecting both the leader sequence and the variable (V) region. Four unique amino acid substitutions were found at positions -8, -3, -2 and -1 in the leader sequence and probably resulted in an unusual cleavage by signal peptidase, thus making the LC truncated by one residue and accounting for its unique hydrophobic N-terminus: Ile-Ile-Leu. Additional peculiarities were observed in the V region, including a Thr74 → Asn substitution creating a N-glycosylation site, and Thr53 → Ile, which was only reported once among human κIII chains, in another LCDD case, and may be of special significance at a position usually harbouring a polar amino acid.
Preferential elimination of CD28+ T cells in systemic lupus erythematosus (SLE) and the relation with activation-induced apoptosis
Authors: Kaneko H , Saito K , Hashimoto H , Yagita H , Okumura K , Azuma M
Published: November 2003

CD28 on T cells provides a potent costimulatory signal for T cell activation. Down-regulation of CD28 on peripheral T cells has been reported in certain clinical conditions, but full studies on the mechanism and biological significance have not been performed. Our extensive phenotype analysis of peripheral blood lymphocytes (PBL) from SLE patients revealed that the absolute number of CD28+ T cells of both CD4 and CD8 phenotypes was selectively decreased, while that of CD28− T cells was maintained. CD28+ T cells from SLE patients exhibited mostly normal proliferative responses to both CD28-dependent and -independent stimulations. In contrast, CD28− T cells were hyporesponsive to anti-CD3 stimulation in both SLE and normal controls. These results implied that the selective decrease of CD28+ T cells in SLE does not result from a hyporesponsiveness of CD28+ T cells. To investigate the reason for the selective loss of CD28+ T cells, we determined the appearance of apoptotic cells in culture with or without anti-CD3 stimulation. Apoptotic cells defined by merocyanine (MC)540 were gradually increased from 12 h to 24 h. Anti-CD3-induced apoptosis of CD28+ T cells was significantly accelerated in SLE, whereas apoptosis of CD28− T cells was hardly detected in both SLE and normal controls. Comparative analysis between CD28+ and CD28− T cells on CD95 (Fas) and Bcl-2 expression, which are related to activation-induced cell death (AICD), did not show a major difference, although CTLA4, which has been demonstrated to transmit an apoptosis-inducing signal, was expressed only on CD28+ T cells. Our results suggest that CD28-mediated costimulation influences T cell susceptibility to AICD and may be involved in T cell lymphopenia in SLE.