bluuebunny/crossref_metadata_2025 huggingface dataset:
Loss of the CD5+ and CD45RAhi B cell subsets in alcoholics
Authors: Cook R T , Waldschmidt T J , Cook B L , Labrecque D R , Mclatchie K
Published: November 2003

Chronic alcoholics are frequently immunodeficient, have polyclonal hypergammaglobulinaemia, and often have autoantibodies. Recent work in other diseases has shown that functional distinctions of possible relevance to autoimmunity and immunodeficiency can be found among the B cell subsets defined by differential expression of the surface markers CD5 and CD45RA. Therefore, we have evaluated the CD5,CD45RA B cell subsets of both chronic alcoholics without evidence of active liver disease (AWLD), and alcoholics admitted for acute alcoholic liver disease (ALD). Mean B cell numbers were normal in AWLD, but significantly reduced in ALD. Analysis of B cells by three-colour flow cytometry in 20 patients and 29 controls revealed a sharp decrease in the percentage of alcoholics’ B cells which were CD5+, 37.6% versus 16.3%, P<0.00001; absolute CD5+ B cell numbers were similarly reduced (58.9 cells/μl versus 20.9; P=0.0012). In addition to the loss of CD5+ B cells, there was a reduction in the percentage of B cells which are CD5−CD45RAhi, leaving many patients with a B cell profile which was predominantly CD19+CD5−CD45RAlo. This subset appears phenotypically similar to the IgM-producing CD5−CD45RAlo subset described by others, and may be enriched for autoantibody-producing cells. One outlier patient was an ALD with 61% of B cells which were CD5+, which also is a profile consistent with increased autoantibody production.
Presence of covalent bonds between immune deposits and other macromolecules in murine renal glomeruli
Authors: Mannik M
Published: November 2003

Microscopic studies have suggested that immune deposits persist for a long time in subepithelial areas of renal glomeruli in human diseases and in experimental models. This study showed quantitatively that in an experimental murine model radiolabelled antibodies in subepithelial immune deposits remained constant for 9 months at about 3 μg/mouse. The extraction of these antibodies was incomplete with 1% SDS at 8 months after immune deposit formation. Furthermore, in the extracted material some antibodies were covalently bound to other molecules. This study demonstrates that covalent bond formation contributes to the persistence of immune deposits in renal glomeruli.
T cell responses to myeloperoxidase (MPO) and proteinase 3 (PR3) in patients with systemic vasculitis
Authors: Griffith M E , Coulthart A , Pusey C D
Published: November 2003

T cell-mediated immune responses are likely to be important in the pathogenesis of systemic vasculitis. However, identifying the T cells involved has proved difficult, and there are conflicting reports regarding T cell proliferation in response to different autoantigens. Perinuclear (P) and cytoplasmic (C) anti-neutrophil cytoplasmic antibodies (ANCA) are closely associated with systemic vasculitis, and are generally specific for MPO or PR3, respectively. We studied the proliferative responses to MPO and PR3 of peripheral blood mononuclear cells from patients with P-ANCA or C-ANCA specific for these antigens by ELISA. These responses were compared with those of normal controls, and of disease controls with P- or C-ANCA not specific for MPO or PR3. The patient group as a whole showed significant T cell proliferation in response to the autoantigens compared with controls (P=0.005). Cells from nine of 13 P-ANCA-positive, anti-MPO-positive patients proliferated in response to MPO, compared with five of 16 controls (P=0.04). Cells from five of eight C-ANCA-positive, anti-PR3-positive patients proliferated in response to PR3, compared with two of 11 controls (P=0.05). These experiments demonstrate that patients with P-ANCA or C-ANCA possess T cells which respond to MPO or PR3, respectively. As in other autoimmune diseases, responses to both antigens were also seen in a proportion of healthy controls. Further analysis of these responses will be important in understanding the pathogenesis of systemic vasculitis and in designing specific immunotherapy.
Cytokine-induced differential expression of serum amyloid A genes in fetal and neonatal rabbits
Authors: Rygg M
Published: November 2003

Serum amyloid A (SAA) is an acute-phase plasma protein which increases 100- to 1000-fold in response to inflammatory stimuli. In this study pregnant rabbits were subjected to laparotomy and their fetuses were injected with lipopolysaccharide (LPS) or various cytokines. Newborn rabbits were likewise stimulated. Hepatic SAA mRNA was studied using Northern blot analyses and scanning densitometry. In vitro derived RNA was used as standard for quantitative mRNA analyses. Cytokine concentrations in amniotic fluid and serum were analysed by biological assays. Fetal rabbits responded to cytokine stimulation by increased hepatic SAA mRNA expression, both during late gestation and in the early neonatal period. However, differences in dose–responses, kinetics and mRNA concentrations were seen dependent on gestational age. IL-1 and tumour necrosis factor (TNF) induced hepatic accumulation of both SAA1, SAA2 and SAA3, while only SAA1 and SAA2 mRNA accumulation was seen after IL-6 stimulation. High levels of IL-1 and TNF found in amniotic fluid from LPS-stimulated fetal rabbits corresponded with high levels in fetal, but not in maternal, serum. High levels of IL-1 and TNF, but no IL-6, were seen in newborn control sera and in adult serum 1 day after a normal delivery. The study details the complexity of the cytokine-induced in vivo response of SAA mRNA in fetal and neonatal rabbits.
Retinoic acid modulates the in vivo and in vitro growth of IL-6 autocrine human myeloma cell lines via induction of apoptosis
Authors: Levy Y , Labaume S , Colombel M , Brouet J C
Published: November 2003

We previously showed that IL-6 is an autocrine growth factor for two human myeloma cell lines, RPMI 8226 and U266. We investigated here the in vitro and in vivo effects of all-trans retinoic acid (RA) on the growth and survival of these two cell lines. RA induced a dramatic dose- and time-dependent inhibition of the proliferation of both cell lines. This inhibition was correlated with a down-modulation of the cell surface expression of the IL-6 binding chain (gp80) and the transducing chain (gp130) of the IL-6 receptor (IL-6R). Long-term culture experiments showed that down-modulation of gp80 expression was complete at days 15 and 30 in the presence of 10–5 and 10–7 mol/l of RA, respectively. Gp130 expression was greatly decreased, albeit still detectable, in similar culture conditions. RA-mediated interruption of the IL-6 autocrine loop was associated with a decrease of bcl-2 oncoprotein expression and apoptosis of the myeloma cells which was RA concentration- and time-dependent. The in vivo relevance of the effects of RA was studied on tumours which developed in nude mice inoculated with a subclone of RPMI 8226. Whereas tumours grew in all control mice, 40% of tumours regressed within 20 days in RA-treated mice. Cells from regressing tumours featured characteristics of apoptosis and exhibited low gp80 and gp 130 expression. Our study indicate that long-term RA treatment interferes in vivo and in vitro with IL-6 autocrine growth of myeloma cell lines, leading to apoptosis.
Differential nitric oxide (NO) production by macrophages from mice and guinea pigs infected with virulent and avirulent Legionella pneumophila serogroup 1
Authors: Rajagopalan-Levasseur P , Lecointe D , Bertrand G , Fay M , Gougerot-Pocidalo M-A
Published: November 2003

l-arginine-dependent reactive nitrogen intermediates have been identified as macrophage cytotoxic effector molecules against intracellular pathogens. To determine its role, ex vivo production of NO by peritoneal macrophages of C3H/HeN mice and Dunkin–Hartley guinea pigs infected intraperitoneally with a virulent and isogenic avirulent Legionella pneumophila serogroup 1 strain was compared with bacterial clearance from the lungs. While the virulent strain was cleared from mice lungs, the guinea pigs died within 96 h. In vivo infection with both strains resulted in the production of NO by mouse peritoneal macrophages ex vivo. In contrast, guinea pig macrophages did not produce detectable NO. In addition, infection by the avirulent strain led to the production of significantly more NO by mouse macrophages than the virulent parent strain, irrespective of stimulation with lipopolysaccharide (LPS) and/or interferon-gamma (IFN-γ). These results suggest that resistance to Leg. pneumophila infection may depend on the production of NO by host macrophages.
Tumour necrosis factor-alpha (TNF-α) gene polymorphism and expression in pre-eclampsia
Authors: Chen G , Wilson R , Wang S H , Zheng H Z , Walker J J , McKillop J H
Published: November 2003

Pre-eclampsia is an endothelial disorder and TNF-α has fundamental effects on endothelial cells by several means, including altering the balance between oxidant and anti-oxidant, changing the pattern of prostaglandin production and affecting expression of several cell surface components. To determine whether TNF-α mRNA expression is increased in pre-eclamptic patients, leucocytes from pre-eclamptic patients, normal pregnant women and normal non-pregnant women were studied for TNF-α expression using a quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method. Using a model of 373A ABI Sequencer, TNF-α gene polymorphism was also analysed by GENESCAN and Genotyper software in order to explain the mechanism of abnormal TNF-α expression. Our results show that TNF-α mRNA expression is significantly elevated in pre-eclamptic patients compared with the other two control groups. The high expression of TNF-α may be associated with the TNF1 allele, whose frequency is markedly increased in pre-eclamptic patients. These observations are consistent with a major role for TNF-α in mediating endothelial disturbances, and suggest a key role for TNF-α in the development of pre-eclampsia.
Defective B7 expression on antigen-presenting cells underlying T cell activation abnormalities in systemic lupus erythematosus (SLE) patients
Authors: García-Cózar F J , Molina I J , Cuadrado M J , Marubayashi M , PeÑa J , Santamaría M
Published: November 2003

Defective T cell functions, including IL-2 production and proliferation, have been shown in SLE patients. After T cell stimulation (first signal), a costimulatory signal (second signal) is required to achieve complete T cell activation. Main costimulatory signals are provided to T cells by B7 antigens (CD80 and CD86, expressed on antigen-presenting cells (APC)) upon interaction with its receptor, the CD28 molecule expressed on T cells. The aim of this study was to investigate the role of CD28/B7 interactions in the impaired T cell responses of SLE patients. We show that stimulation of T cells with phytohaemagglutinin (PHA) in the presence, but not in the absence, of anti-CD28 MoAb or B7+ cells results in tyrosine phosphorylation of specific substrates, transcription of mRNA and production of IL-2 that is indistinguishable in SLE patients and healthy controls. Moreover, proliferation of costimulated T cells from SLE and controls was specifically abrogated by blocking the CD28/B7 interactions by means of addition to the culture of the CTLA4-Ig fusion protein. However, in most patients activated APC failed to up-regulate B7 molecules, giving rise to ineffective costimulatory signalling to T cells. These results indicate that the CD28/B7 costimulatory pathway is defective in SLE patients.
Anti-neutrophil cytoplasmic antibodies (ANCA) directed against bactericidal/permeability increasing protein (BPI): a new seromarker for inflammatory bowel disease and associated disorders
Authors: Stoffel M P , Csernok E , Herzberg C , Johnston T , Carroll S F , Gross W L
Published: November 2003

It was our purpose to determine the immunodiagnostic value of ANCA directed against BPI in diseases known to be associated with ANCA, such as ANCA-associated vasculitides, inflammatory bowel disease (IBD) and the associated condition primary sclerosing cholangitis. The immunoreactivity of recombinant BPI (rBPI) was established in order to develop an ELISA specific for rBPI. By means of this assay, BPI-ANCA were assessed in sera of 178 patients with IBD or the associated disorder primary sclerosing cholangitis, 112 patients with ANCA-associated vasculitides, and in sera of 182 disease and 140 healthy controls. BPI-ANCA were found to be closely associated with IBD and primary sclerosing cholangitis (34% and 44% of ANCA-positive sera, respectively). By contrast, BPI-ANCA positivity was low (<10%) in the double-negative sera of patients with ANCA-associated vasculitides and in disease and healthy controls. BPI-ANCA appear to constitute an important marker for IBD and primary sclerosing cholangitis, but not for the ANCA-associated vasculitides.
Prevention of adjuvant arthritis in Lewis rats by neonatal bacille Calmette–Guérin (BCG) infection
Authors: Esaguy N , Águas A P
Published: November 2003

Tolerization of pathogenic antigens is one of the experimental strategies that has been proposed to prevent autoimmune disease. We have investigated here whether neonatal intraperitoneal infection of Lewis rats with Mycobacterium bovis-BCG has any effect on the expression of adjuvant arthritis (AA), an autoimmune disease that is produced by immunization of the rats with dead mycobacteria in mineral oil (i.e. Freund’s complete adjuvant (FCA)). We found that neonatal infection with 108 viable BCG bacilli rendered all Lewis rats resistant to the expression of AA after FCA immunization. This BCG-induced protection from reactive arthritis was not seen in Lewis rats infected with smaller inocula (106 BCG bacilli) or if the infection was performed after the neonatal period (e.g. at 3 weeks of age). Neonatal administration of 65-kD mycobacterial heat shock protein (hsp65, a key antigen in the etiopathogenesis of AA) failed to protect Lewis rats from AA; injection of lactoferrin (an autoantigen that may be involved in the physiopathology of autoimmune arthritis) to newborn Lewis rats decreased the severity of AA observed after FCA immunization of the animals. Western blotting revealed that Lewis rats that had acquired resistance to AA also showed changes in their repertoire of antibody specificities; among these alterations was decreased anti-hsp65 reactivity. We conclude that neonatal infection with BCG, but not hsp65 injection, renders Lewis rats resistant to AA and that the phenomenon is associated with change in the repertoire of specificities of circulating antibodies.
Longitudinal study of the frequency of cytotoxic T cell precursors in kidney allograft recipients
Authors: Mestre M , Massip E , Bas J , Alsina J , Romeu A , Castelao A M , Buendia E , Grinyó J M
Published: November 2003

Clonal deletion or inactivation of donor-specific alloreactive cells are important mechanisms that are believed to account for acquired immune tolerance in allograft recipients. Serial assessment of precursor cytotoxic T lymphocyte frequencies (CTLpf) by limiting dilution analysis (LDA) provides information at the clonal level on changes in the alloimmune response of graft recipients. We performed a longitudinal study of 15 cadaveric kidney recipients before and every 3 months throughout the first year after transplantation (Tx). Pre-Tx values of donor CTLpf showed high interindividual variability without a predictive value for the clinical outcome. All patients with well functioning kidneys had decreased CTLpf at 3 months post-Tx in comparison with pre-Tx values. This decrease was donor-specific in four patients and was permanent in two cases throughout the study. Most patients presented decreased anti-donor CTLpf values from 6 to 9 months, whereas a partial recovery of donor CTLpf was observed in three patients. Reversible acute rejection was diagnosed in three patients, and it was associated with a marked increase in anti-donor CTLpf, returning to pre-Tx values by 9 months post-Tx. In addition, one patient with chronic rejection displayed a transient increase in CTLpf 6 months after Tx. The results of this sequential study indicate the establishment of a state of either hyporesponsiveness or functional clonal inactivation, transient or permanent, which could facilitate allograft acceptance.
Induction of autoimmune hepatitis and autoantibodies to liver antigens by neonatal thymectomy in mice
Authors: Myozaki M , Kamiyasu M , Miura T , Watanabe Y , Nakanishi T , Yamashita U
Published: November 2003

We examined development of autoimmune hepatitis in neonatally thymectomized C3H/HeN mice and tried to characterize the nature of liver antigens recognized by the autoantibodies at the molecular level. Autoantibodies to crude liver proteins detected by ELISA were found in 12 (67%) of 18 mice thymectomized 2 days after birth. However, autoantibodies were not detected in mice thymectomized 7 days after birth. The autoantibodies mainly consisted of IgG and reached the maximum level 8 weeks after birth. Hepatic inflammation, mononuclear cell infiltration in the portal area, was seen in 5 (28%) of 18 mice thymectomized 2 days after birth, but not in mice thymectomized 7 days after birth. Most infiltrating cells were Thy-1+ lymphocytes. The serum autoantibody level to crude liver proteins in mice with hepatitis was much higher than that in mice without hepatitis. We fractionated crude liver proteins by a Sepharose 6B column and examined the reactivity against the autoantibodies. The autoantibodies of three of five mice with hepatitis reacted with the ≈150 kD liver proteins other than liver-specific protein (LSP). By Western immunoblotting of SDS–PAGE using LSP and fractionated liver proteins, we found that the molecular weights of the target antigens were 52 kD in LSP and 150 kD (strong band), 138, 128, 120 and 110 kD (weak band) in fractionated liver proteins other than LSP. This 150-kD target molecule in crude liver proteins was found only in liver. These results indicate that hepatitis and autoantibodies to liver proteins are induced spontaneously by neonatal thymectomy in mice, and the candidates of autoantigen in this hepatitis model are 52-kD protein in LSP and 150-kD liver proteins different from LSP. Still more, we regard the 150-kD molecule as a new autoantigen related to hepatitis.
Transforming growth factor-beta (TGF-β) expression and interaction with proteinase 3 (PR3) in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis
Authors: Csernok E , Szymkowiak C H , Mistry N , Daha M R , Gross W L , Kekow J
Published: November 2003

TGF-β is a multifunctional cytokine modulating the onset and course of autoimmune diseases as shown in experimental models. The aim of this study was to investigate TGF-β expression in ANCA-associated vasculitis (AAV), and the possible interactions of this cytokine with lysosomal enzymes identified as ANCA autoantigens (e.g. PR3). This included TGF-β effects on the translocation of the lysosomal enzymes to the cell surface of polymorphonuclear neutrophils (PMN), and the presumed activation of non-bioactive, latent TGF-β by these enzymes. Patients with various types of systemic vasculitis (SV) were studied, including three different types of AAV (Wegener's granulomatosis (WG), Churg–Strauß syndrome (CSS) and microscopic polyangiitis (MPA)). Regardless of the type of assay applied, the TGF-β1 isoform was found to be overexpressed in SV, including AAV, and to correlate with disease activity as shown for WG. Mean TGF-β1 plasma levels in AAV patients ranged from 8.9 ng/ml (WG) to 13.3 ng/ml (CSS) (control 4.2 ng/ml; P < 0.01), while TGF-β2 levels were not elevated. Flow cytometry analysis showed TGF-β1 to be a potent translocation factor for PR3 comparable to other neutrophil-activating factors such as IL-8. PR3 membrane expression on primed PMN increased by up to 51% after incubation with TGF-β1. PR3 itself was revealed as a potent activator of latent TGF-β, thus mediating bioeffects of this cytokine. These findings, together with other features of TGF-β such as induction of angiogenesis and its strong chemotactic capacity, indicate that TGF-β might serve as a proinflammatory factor in SV, especially in AAV.
Activation antigen expression on human T cells. I. Analysis by two-colour flow cytometry of umbilical cord blood, adult blood and lymphoid tissue
Authors: Amlot P L , Tahami F , Chinn D , Rawlings E
Published: November 2003

Activation antigens (actags) were detected on T cells at low levels of intensity by carefully defining negative cells with a panel of control antibodies. The mean percentage of blood T cells from healthy volunteers that expressed actags were 22% (CD25), 54% (CD26), 38% (CD38), 12% (CD54), 6% (CD69) and 21% (HLA-DR). The variability of actag expression detected by this sensitive method was determined on healthy volunteers by repeated estimation over a year. The percentage of T cells expressing CD25 and CD26 varied no more than repeated estimation of the CD4 T cell subset, whereas other actags showed greater variability. The antigen density of these actags on T cells was determined in relation to CD4 antigen density, and for most actags ranged from 10% to 75% of the level of CD4 antigen density except for CD7 and HLA-DR, which could exceed that of CD4. Different degrees of actag expression characterized T cells from different blood and lymphoid tissues. CD26, CD38 and CD45RA were universally expressed in cord blood at higher antigen density than adult blood. This immature pattern was consistent with recent thymic emigration. CD25, CD45RO, CD54 and HLA-DR progressively increased from cord blood through adult blood to lymphoid tissues, consistent with antigen-driven activation, whereas CD26 and CD45RA decreased. CD69, a very early activation antigen, abruptly increased in lymphoid tissue, exceeding CD25 by two-to-three-fold and suggesting a pre-activation state that may not involve commitment to antigen-driven proliferation. CD7 and CD38 expression was higher in cord blood and lymphoid tissue than in adult blood, indicating both an antigen-independent and -dependent up-regulation.
Tyrosinase as an autoantigen in patients with vitiligo
Authors: Baharav E , Merimski O , Shoenfeld Y , Zigelman R , Gilbrud B , Yecheskel G , Youinou P , Fishman P
Published: November 2003

Vitiligo is considered an autoimmune disorder due to the generation and presence of autoantibodies directed against melanocyte antigens in the patients' sera. In the present study we point towards a newly defined autoantigen in vitiligo, the enzyme tyrosinase, which participates in the process of melanogenesis. Anti-tyrosinase antibodies were detected in the sera of seven patients with diffuse and 11 patients with localized vitiligo. Employing solid-phase ELISA to mushroom tyrosinase, we found that patients with diffuse vitiligo had significantly higher titres of IgG anti-tyrosinase autoantibodies than patients with localized disease or healthy subjects. These anti-tyrosinase autoantibodies have relatively high functional affinity to tyrosinase and can be recovered from vitiligo patients’ sera by affinity purification. The anti-tyrosinase antibodies do not cross-react with other enzymes recognized as autoantigens in different autoimmune disorders and the autoantibodies do not block the enzymatic activity of tyrosinase, indicating that they are not reacting with the catalytic site of the enzyme. These data point to tyrosinase as an autoantigen in vitiligo and suggest that anti-tyrosinase titres can serve as a marker for disease activity.
Complement-dependent cytotoxic antibodies to human T lymphotropic virus type I (HTLV-I)-infected cells in the sera of HTLV-I-infected individuals
Authors: Plotni Cky-Gilquin H , Afani A , Vernant J C , Desgranges C
Published: November 2003

To investigate whether HTLV-I induces the development of complement-dependent cytotoxic antibodies in humans, sera of asymptomatic HTLV-I carriers and of patients suffering from tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM) or adult T cell leukaemia (ATL) were used in a cytotoxicity assay against a panel of target cells. This panel included uninfected cell lines (CEM, Jurkat, Molt and H9), cell lines chronically infected with HTLV-I (MT2, MT4, C91PL and HUT102), as well as lines H36 (H9 infected with HTLV-I), H9-IIIB (H9 infected with HIVIIIB) and H9-MN (H9 infected with HIVMN). HTLV-I+ sera induced lysis of H36 and of lines expressing HTLV-I antigens in the presence of rabbit complement, but did not lyse cells in presence of human complement. The HTLV-I+ sera also failed to lyse the HTLV-I− lines and H9 cells, suggesting that lysis was specific for HTLV-I. H36 cell lysis was prevented by IgG depletion of the sera and by dialysis of rabbit complement against EGTA or EDTA. Rabbit complement-dependent cytotoxic antibodies were present in the sera of 14/14 HTLV-I-infected individuals; the highest titres were predominantly found in the sera of the TSP/HAM patients. Such antibodies were also detected in 5/5 individuals coinfected with HIV-1 and HTLV-I, although no cytotoxic antibody could be found against HIV-infected cells. Vice versa, sera of HIV-1-infected individuals did not exert a lytic effect in the presence of complement (of human or rabbit origin) against HIV-1- or HTLV-I-infected cells. Incubation of the sera of four HTLV-I-infected patients with HTLV-I env-specific synthetic peptides demonstrated that some of the complement-dependent cytotoxic antibodies recognized epitopes located on gp46 between amino acids 190 and 209. There is no correlation of rabbit complement-dependent cytotoxic HTLV-I antibodies with the development of disease.
Autoantibodies to thyroid hormones: the role of thyroglobulin
Authors: Erre Gragui K , Cheillan F , Defoort J P , Prato S , Fert V
Published: November 2003

Autoantibodies against thyroid hormones (THAA) are frequently detected in the sera of patients with thyroid disorders together with autoantibodies against thyroglobulin (TGAA). THAA are considered to be a subset of TGAA, but alternative possibilities have not been excluded. We hypothesize that if THAA arise through an immune response to iodothyronines carried by circulating thyroglobulin (hTg), THAA should be found together with autoantibodies against the peptide backbone of hTg (TPAA) close to the hormone-forming sites. We measured TPAA in 178 serum samples, obtained from healthy subjects and patients with thyroid disorders, using two hormone-forming peptides isolated from hTg. The occurrence of TPAA was much lower than that of TGAA. Autoantibodies to the hormone-rich peptide, P3, were significantly more common than autoantibodies to the hormone-poor peptide, P1 (111/178 = 62.3% for TGAA versus 21/178 = 11.8% for anti-P3 TPAA and 7/178 = 3.9% for anti-P1 TPAA). The presence of autoantibodies to thyroid hormones was investigated in 25 TPAA+ and 26 TPAA− sera. THAA were found more frequently in TPAA+ sera (10/25 = 40% for TPAA+ and 4/26 = 15.3% for TPAA−). Correlation analysis shows that the anti-P3, but not the anti-P1 binding activity, correlates positively with the THAA-binding activity (P < 0.001 for anti-T4 THAA; P < 0.01 for anti-T3 THAA). Specificity of anti-P3 TPAA indicates that a subset of the anti-P3 antibodies is directed against the thyroid hormone moiety and another subset is directed against the peptide backbone near the hormone-forming peptide, according to our hypothesis. These results indicate that the THAA response is an anti-hTg response directed, in a significant number of cases, against the hormone-forming site included in the P3 peptide. This response seems to be elicited by either native hormone-rich hTg or by hTg fragments.
The mononuclear phagocyte–dendritic cell dichotomy: myths, facts, and a revised concept
Authors: Goerdt S , Kodelja V , Schmuth M , Orfanos C E , Sorg C
Published: November 2003

Since Aschoff's reticuloendothelial system was abandoned a few decades ago, classification and characterization of the mononuclear phagocyte and dendritic cell systems have evolved separately or even in competition with one another. New information has now become available indicating that monocytes/macrophages and dendritic cells have a common origin in the bone marrow, and may even transdifferentiate. Morphological and functional distinctions—although valid under certain conditions—have been blurred by revelation of the versatility of monocytes/macrophages and dendritic cells in response to different contextual needs in inflammation and immunity. Monocytes/macrophages and dendritic cells share a sentinel, receptor/effector, and presentation mode, and may either activate or silence specific immune reactions. In keeping with the view of monocytes/macrophages and dendritic cells as interactive sentinels, we suggest that the mononuclear phagocyte and dendritic cell systems be replaced by the custocyte system (custos, Lat = sentinel, guard) as a unifying concept. Within the custocyte system, we recognize type I, type II, and type III custocytes. Type I and II custocytes exhibit predominance of presentation or effector/presenter interdependency, respectively, while type III custocytes are bipolar, passing through type I- and type II-like phases during their development and in inflammatory responses. The custocyte system brings into view monocytes/macrophages and dendritic cells as dynamic players in immunity and inflammation with a high degree of derivational, phenotypic, functional, and molecular plasticity.
Helicobacter pylori infections in IgA deficiency: lack of role for the secretory immune system
Authors: Bogstedt A K , Nava S , Wadström T , Hammarström L
Published: November 2003

During the last years considerable attention has been focused on the possibility of the development of an oral vaccine against Helicobacter pylori. However, Helicobacter infection is known to be life-long, despite a vigorous immune response, and the hypothesis that an increased local production of secretory IgA in the gastric mucosa, due to vaccination, should protect against the colonization of the bacterium may therefore be questioned. In this study, when comparing the seropositivity and titre against H. pylori in IgA-deficient patients and age-related normal blood donors, it appears that lack of secretory IgA does not seem to have any major influence on the prevalence of the infection, nor is it reflected in titres of specific IgG antibodies. These results may argue against a pivotal role for IgA in the defence against Helicobacter, and raise questions about current strategies for the development of an oral vaccine against H. pylori and may point to a need for alternative therapeutic strategies.
CD8+ lymphocyte phenotype and cytokine production in long-term non-progressor and in progressor patients with HIV-1 infection
Authors: Zanussi S , Simonelli C , D'Andrea M , Caffau C , Clerici M , Tirelli U , De Paoli P
Published: November 2003

In most HIV-1-infected patients, clinical and immunological progression develops within a few years. Few infected people, termed long-term non-progressors (LTNP), remain healthy and immunologically stable for a long time. The factors governing the maintenance of this condition are not well known, but it is conceivable that CD8+ lymphocytes, cells that play a central role in controlling in vitro HIV replication, may have a part in vivo in this process. The aim of this study was to characterize the phenotypic profile and the cytokine production of CD8+ cells in a group of LTNP patients who had stable CD4+ cell counts (>500/mm3) for at least 7 years. Their CD8+ absolute numbers were similar to a control group composed of HIV-1+ patients who have a progressive decline of their CD4+ cell counts. However, our multiparameter immunofluorescence studies show that a clinical and immunologically stable condition is associated with the presence of a CD28+, CD95 strongly positive CD8+ population, while disease progression is marked by the CD28−CD95+CD8+ subset. Purified CD8+ cells from LTNP retain their ability to produce IL-2, interferon-gamma (IFN-γ) and, to a lesser degree, to produce IL-10 and IL-4. In contrast, CD8+ cells from progressors are unable to secrete IL-2 and IL-10. Although CD8+ cytokine profile does not fit with the proposed T helper (Th)1/Th2 switch in progressive HIV infection, LTNP CD8+ T cells maintain their capacity to produce IL-2 and IL-10 (Th0-like), a pattern very similar to that observed in normal HIV healthy controls. We suggest that CD8+ cells expressing CD28, CD95 and having a Th0-like profile may be considered to be associated with long-term survival.
Immunoregulation in onchocerciasis: predominance of Th1-type responsiveness to low molecular weight antigens of Onchocerca volvulus in exposed individuals without microfilaridermia and clinical disease
Authors: LÜder C G K , Schulz-Key H , Banla M , Pritze S , Soboslay P T
Published: November 2003

Chronic and generalized onchocerciasis is associated with suppression of the parasite-specific cellular responsiveness, while exposed individuals without parasitological and clinical evidence of infection (endemic normals) display prominent cellular reactivity to Onchocerca volvulus antigens (OvAg). In order to identify those parasite antigens which may account for this differential cellular responsiveness, total adult worm-derived OvAg were fractionated by means of preparative SDS–PAGE and blot-elution into 22 antigen fractions of continuously decreasing molecular weight. Peripheral blood mononuclear cells (PBMC) from microfilariae (mf)-positive onchocerciasis patients (n = 18) proliferated weakly in response to all OvAg fractions. In contrast, in vitro reactivity of PBMC from endemic normals (n = 9) was depressed in response to OvAg of mol. wt 200–30 kD only, while antigens of mol. wt <30 kD induced vigorous proliferation in these individuals compared with the microfilaridermic patients (P < 0.05). Highest proliferative reactivity of cells from endemic normals was observed in response to OvAg of mol. wt 15–11 kD. Furthermore, these low mol. wt antigen fractions induced substantial production of IL-2 and interferon-gamma (IFN-γ) in PBMC from endemic normals, but not in those from onchocerciasis patients. Cells from individuals of both groups secreted similar amounts of IL-5 in response to all OvAg fractions, with highest production again being induced by low mol. wt OvAg. In contrast, PBMC from onchocerciasis patients clearly produced more IL-10 than did cells from endemic normals. This augmented IL-10 production by PBMC from mf-positive individuals was not only observed after stimulation with OvAg fractions, but was measured in unstimulated control cultures as well. IFN-γ-specific mRNA in antigen-stimulated PBMC from endemic normals appeared to be more prominent than in cells from onchocerciasis patients. However, mRNA transcripts of IL-10 and IL-13 were clearly present in patients, but were absent or inconsistently observed in endemic normals. Our results suggest that vigorous Th1-type cellular responsiveness encountered in endemic normals is restricted to low mol. wt antigens of O. volvulus, while such reactivity will not be present in mf-positive individuals. Furthermore, spontaneous production of high levels of IL-10 in onchocerciasis patients is likely to suppress Th1-type immunity, and thus may favour manifestation of chronic onchocerciasis. These traits of cellular immunity may contribute to the differential outcome of O. volvulus infection, the manifestation of clinical disease, and may also regulate the build up of acquired immunity in humans.
Immunization of non-obese diabetic (NOD) mice with glutamic acid decarboxylase-derived peptide 524–543 reduces cyclophosphamide-accelerated diabetes
Authors: Saï P , Rivereau A S , Granier C , Haertlé TH , Martignat L
Published: November 2003

NOD mice constitute a model for studying the prevention of human autoimmune type 1 diabetes. Glutamic acid decarboxylase (GAD) could be a key antigen involved in this disease, and GAD65 peptide 524–543 has been implicated in early T cell response in young NOD mice. We performed two i.p. injections of GAD peptide 524–543 (100 μg at each injection), together with Freund's incomplete adjuvant (FIA), into female NOD mice at 30 and 45 days old. Diabetes was accelerated 2 weeks later by a single injection of cyclophosphamide (CY), which acts against suppressive mechanisms. Treatment with GAD 524–543 peptide delayed the onset of diabetes and reduced its incidence (28% versus 60%; P<0.001) compared with control mice injected with FIA alone, or GAD peptide 534–553, or an irrelevant peptide. In the same group, the severity of lymphocytic inflammation of pancreatic islets was reduced (P<0.03). Up to 3 months after peptide injections, a strong splenocytic proliferative response occurred in immunized NOD mice against the immunizing peptide alone (but not against a panel of seven other GAD65-derived peptides). After peptide challenge of splenocytes in vitro, protection against CY-accelerated diabetes was associated with higher peptide-specific production of T helper type 2 (Th2)-associated interleukins 4 and 10, whereas Th1-associated interferon-gamma and IL-2 were proportionally less represented. During cotransfer, T splenocytes from GAD 524–543-immunized mice were able to reduce the capacity of T cells from diabetic donors to transfer the disease adoptively (P<0.01), demonstrating the generation of cellular mechanisms that actively suppress the disease. It is concluded that immunization of NOD mice with GAD65 peptide 524–543 can counteract CY-accelerated diabetes, possibly through active cellular suppression linked to a shift of Th1/Th2 balance toward the production of Th2 cytokines such as IL-4 and IL-10. This study provides additional support for the notion that GAD, and more precisely its epitope 524–543, could be one of the key targets for the pathogenesis of type 1 diabetes in NOD mice, as well as for the efficacy of disease-specific peptide therapy in type 1 diabetes.
Predominance of a type 2 intratumoural immune response in fresh tumour-infiltrating lymphocytes from human gliomas
Authors: Roussel E , Gingras M C , Grimm E A , Bruner J M , Moser R P
Published: November 2003

Increasing evidence suggests the existence of polarized human T cell responses described as Th1-type (promoting cell-mediated immunity) and Th2-type (promoting humoral immunity), characterized by a dominant production of either interferon-gamma (IFN-γ) or IL-4, respectively. Little is known about the intratumoural activation of infiltrating lymphocytes (TIL) in human gliomas. Therefore, we assessed fresh TIL at cellular and molecular levels to find out if they were activated and polarized into a type 1 or 2 immune response. Flow cytometry analysis of TIL revealed that the major subset was made of T lymphocytes. Double labelling with α-CD3 and adhesion/activation markers revealed T cell subsets expressing CD49a, CD49b, CD54, and CD15, some of which were almost absent in autologous T peripheral blood lymphocytes (T-PBL). Furthermore, the proportions of T-TIL expressing CD56, CD65, or CD25 were several-fold higher than in T-PBL. Intratumoural functional activation of TIL was tested by semiquantitative assessment in relative units (RU) of lymphokine gene activation with mRNA reverse transcriptase-polymerase chain reaction (RT-PCR). All TIL populations except one significantly expressed IL-4 1 to 2 logs of RU above healthy PBL baseline. Similarly, all patients expressed granulocyte-macrophage colony-stimulating factor (GM-CSF) in a range comparable to IL-4. However, most TIL populations did not express IFN-γ, IL-2, and tumour necrosis factor-beta (TNF-β) at higher levels than healthy normal PBL. The increased proportion of T cells expressing activation markers and the consistent detection of significant IL-4 and GM-CSF lymphokine gene activation in TIL populations suggested a predominant type 2 intratumoural immune response that does not promote cell-mediated tumouricidal activity and may contribute to the inefficiency of the antiglioma immune response.
Autoantibodies to mitochondrial 2-oxo-acid dehydrogenase complexes in localized scleroderma
Authors: Fujimoto M , Sato S , Ihn H , Tamaki T , Kikuchi K , Soma Y , Tamaki K
Published: November 2003

Sera from patients with localized scleroderma frequently produce cytoplasmic staining by indirect immunofluorescence, although the antigen remains to be determined. We studied the prevalence, antigen specificity and associated clinical characteristics of anti-cytoplasmic antibodies in localized scleroderma. Serum samples from 60 patients with localized scleroderma were examined by indirect immunofluorescence analysis and immunoblotting. By immunofluorescence analysis on HEp-2 cell substrate, seven of 60 (12%) patients were shown to be positive for anti-cytoplasmic antibodies. Among these, six patients with generalized morphea had anti-mitochondrial antibodies as shown by immunoblotting: they showed reactivity with the E2 component of pyruvate dehydrogenase complex (PDC), with protein X, and with the E2 component of α-oxo-glutarate dehydrogenase complex, while two of them showed reactivity with PDC-E1α. One of these patients who was positive for anti-PDC-E1α antibody showed laboratory abnormalities, suggesting the presence of primary biliary cirrhosis. The age of disease onset was significantly higher in these six patients than in those without anti-mitochondrial antibodies. Furthermore, five of them were classified into generalized morphea with multiple plaque lesions but without linear lesions (multiple plaque type). These observations suggest that major antigens for anti-cytoplasmic antibodies in patients with localized scleroderma are mitochondrial enzymes, 2-oxo-acid dehydrogenase complexes. Patients with anti-mitochondrial antibodies may comprise a unique subset of localized scleroderma designated multiple plaque type of generalized morphea of older onset.
Yersinia-hsp60-reactive T cells are efficiently stimulated by peptides of 12 and 13 amino acid residues in a MHC class II (I-Ab)-restricted manner
Authors: Noll A , Autenrieth I B
Published: November 2003

Heat shock proteins (hsp) are immunodominant antigens in microbial infections. Previous work from this laboratory demonstrated that Yersinia-hsp60 (Y-hsp60)-reactive CD4+αβ T cells play an important role for resolution of Y. enterocolitica infections in mice. In the present study we identified two epitopes of Y-hsp60 recognized by CD4+ Th1 cell clones. The epitopes comprise 12 (214–225) and 13 (74–86) amino acid (aa) residues of Y-hsp60, and are the first described for MHC class II (I-Ab) molecules. Both epitopes are also recognized by T cells isolated from mesenteric lymph nodes from mice orogastrically infected with yersiniae. Stimulation of T cells with peptides of 12 and 13 aa residues of Y-hsp60 caused highly efficient proliferation compared with longer peptides, full-length recombinant Y-hsp60, or heat-killed Yersinia (HKY). Incubation of antigen-presenting cells with chloroquine blocked both peptide and HKY-triggered T cell proliferation, whereas cytochalasin B only blocked HKY-induced proliferation and to a lesser extent peptide-induced proliferation. The identified epitopes reside in a region of Y-hsp60 that is conserved between Enterobacteriaceae but highly variable when compared with murine or human hsp60. Although both epitopes are identical to the related sequence of hsp60 (GroEL) of Escherichia coli, only weak T cell responses were observed upon stimulation with GroEL of E. coli, suggesting that other factors, e.g. flanking amino acid residues, might be important for antigen processing and T cell stimulation in a class II-restricted manner. Furthermore, these observations might be of significance for the rational design of subunit vaccines.
Lysis of pulmonary fibroblasts by lymphokine (IL-2)-activated killer cells—a mechanism affecting the human lung microenvironment?
Authors: Zambello R , Trentin L , Enthammer C , Cipriani A , Agostini C , Semenzato G
Published: November 2003

In this study we investigated whether IL-2-activated killer cells may bind and exert lytic activity against non-transformed lung fibroblasts. We demonstrated that human lymphokine-activated killer (LAK) cells generated in vitro following incubation with recombinant IL-2 of either peripheral blood mononuclear cells (PB-LAK) or lymphocytes obtained from bronchoalveolar lavage (BAL-LAK), but not resting cells, can lyse normal lung fibroblasts obtained from transbronchial lung biopsies in a 4-h 51Cr release assay. Both autologous and allogeneic fibroblasts were consistently lysed by LAK cells, thus suggesting that the phenomenon we observed is not MHC-restricted. Since fibroblasts can bind IL-2 through specific receptors, we evaluated whether long-term culture with rIL-2 could modulate the susceptibility to lysis of target cells. Our data showed that autologous fibroblasts were more resistant to lysis than allogeneic fibroblasts when they were cultured with rIL-2. Since LAK cells have been demonstrated to release a series of different immunomodulatory cytokines, we evaluated the effect of short-term incubation of fibroblasts with different factors, including IL-1, IL-2, IL-3, IL-4, IL-6, tumour necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ), on the binding and the lysis mediated by LAK cells. These cytokines were not directly cytotoxic on fibroblasts. Only IFN-γ was found to have a significant protective effect against the lysis. Our data support the concept that a self-directed cytotoxicity against pulmonary fibroblasts is generated during lymphocyte activation with rIL-2.
Synergistic effect of deoxyspergualin (DSP) and cyclosporin A (CsA) in the prevention of spontaneous autoimmune diabetes in BB rats
Authors: Di Marco R , Zaccone P , Magro G , Grasso S , Lunetta M , Barcellini W , Nicolosi V M , Meroni P L , Nicoletti F
Published: November 2003

Dose-dependent side effects are frequently observed with immunosuppressive drugs of potential relevance for the immunotherapy of insulin-dependent diabetes mellitus (IDDM), such as CsA and DSP. If CsA and DSP acted synergistically in vivo, their combined use would allow using each compound at lower doses than those required when each drug is given in monotherapy. Consequently, dose-dependent side effects could be reduced and the therapeutic activity maintained or even enforced. Toward this end we studied the effects of combined treatment with CsA and DSP on the course of IDDM in the diabetes-prone (DP)-BB rat. The results show that two ‘low’ doses of CsA (2mg/kg) and DSP (1mg/kg) that are clinically ineffective in suppressing IDDM development in BB rats when administered alone under a prolonged prophylactic regimen (30–105 days old), may successfully prevent, but not cure, the disease when given contemporaneously under the same experimental conditions. The combined treatment was well tolerated, and no side effects were noticed. These data suggest that the combined use of CsA and DSP may deserve consideration for its possible application in the prevention/treatment of human IDDM and other autoimmune diseases.