bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of extraocular muscle-infiltrating T cells in thyroid-associated ophthalmopathy (TAO)
Authors: PAPPA A , CALDER V , AJJAN R , FELLS P , LUDGATE M , WEETMAN A P , LIGHTMAN S
Published: November 2003

TAO is characterized by an autoimmune process affecting the orbital contents. T cells have been suggested to have a major role in pathogenesis, but so far only limited data are available to clarify the extraocular muscle (EOM)-infiltrating T cell phenotype, antigenic reactivity and cytokine profile in TAO patients. In the present study, biopsies of affected EOM were taken and the infiltrating T cells isolated and expanded in vitro with mitogen. Their phenotype was determined by flow cytometric (FACS) analysis and compared with peripheral blood-derived T cell lines, treated in the same way from the same patient. Cytokines present in the supernatant after mitogen stimulation of the T cell lines were assayed by ELISA. In addition, cytokine mRNA present at the time of biopsy was determined by rapid RNA extraction from EOM and reverse transcription-amplification with specific cytokine oligonucleotide probes (IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-13, IL-15, interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α)). In the T cell lines from two patients, proliferation assays were carried out with antigens derived from thyroid gland, EOM and a thyrotropin (TSH) receptor preparation. Most T cell lines were CD4+, CD45RO+, and TCR α/β+, both from the EOM and the peripheral blood. A wide variety of cytokines was detected by analysis of supernatants or mRNA, but the profiles were not identical comparing the two approaches. However, IL-4 was detected by both. Dose-dependent proliferation was observed in response to thyroid extract in a biopsy-derived T cell line. In conclusion, EOM-infiltrating T cells from patients with TAO, expanded in vitro, were chiefly CD4+ and produced a mixture of cytokines, including IL-4. The proliferation data suggest that there are thyroid-reactive T cells in EOM.
Cytokine responses to the native and recombinant forms of the major surface glycoprotein of Pneumocystis carinii
Authors: THEUS S A , SMULIAN A G , SULLIVAN D W , WALZER P D
Published: November 2003

Pneumocystis carinii is a major opportunistic pathogen and leading cause of morbidity in patients with AIDS. The major surface glycoprotein (MSG) of P. carinii, represented by a family of related proteins encoded by unique genes, is highly immunogenic and contains T cell-protective epitopes. We undertook the present study to define the CD4 T helper (Th) response by cytokine secretion to native MSG and a recombinant form of the protein, MSG-B. Spleen cells were collected from Lewis rats and restimulated with both native MSG and MSG-B. Within 24 h, the CD4 cells secreted high levels of interferon-gamma (IFN-γ) in response to both types of antigen, indicative of a Th1 response; however, after 72 h of incubation, only the native MSG stimulated secretion of IL-4 (Th2 response) from the cells. We then investigated whether the presence of IL-4 could alter the predominant Th1 phenotype by the CD4 cells in response to MSG and MSG-B. Cells cultured with native MSG and IL-4 produced low levels of IFN-γ and elevated levels of IL-4. Interestingly, cells incubated with MSG-B and IL-4 reduced production of IFN-γ, but were not stimulated to produce increased levels of IL-4. The presence of anti-IFN-γ antibody in the MSG- or MSG-B-stimulated cultures did not effect the expression of IFN-γ mRNA, suggesting that the generation of Th1 cells in response to MSG or MSG-B was not dependent on IFN-γ. We conclude that native MSG, which contains multiple forms of this antigen, and recombinant MSG elicit different cytokine responses in vitro. These data are not only important to studies of MSG, but may also be relevant to the role of MSG in the immunopathogenesis of P.carinii infection in vivo.
Human peritoneal B-1 cells and the influence of continuous ambulatory peritoneal dialysis on peritoneal and peripheral blood mononuclear cell (PBMC) composition and immunoglobulin levels
Authors: DONZE H H , LUE C , JULIAN B A , KUTTEH W H , KANTELE A , MESTECKY J
Published: November 2003

In mice, peritoneal B cells are composed of a unique B-1 cell population which can repopulate the intestinal lamina propria with IgA-producing cells, as well as contribute to the majority of serum IgM. In this study, peritoneal lymphocytes from patients starting continuous ambulatory peritoneal dialysis (CAPD) and from women undergoing bilateral tubal ligation (BTL) were analysed for the presence of a B-1 cell population as well as the expression of potential homing receptors. Up to 63% of the peritoneal B cells express surface antigen CD5, and most peritoneal lymphocytes express the mucosal homing receptors, α4β7 and αEβ7. When analysing serial samples collected from patients from the beginning of dialysis to 1 year, no marked changes were observed in serum or salivary immunoglobulin levels, although the peritoneal lymphocyte population was reduced by 50%. These data suggest that the phenotype of human peritoneal B-1 cells is similar to that of mice, but the contributions to the immune system may differ.
Oral tolerance in a murine model of relapsing experimental autoimmune uveoretinitis (EAU): induction of protective tolerance in primed animals
Authors: THURAU S R , CHAN C-C , NUSSENBLATT R B , CASPI R R
Published: November 2003

Oral administration of uveitogenic retinal antigens suppresses the expression of EAU induced by a subsequent immunization with these antigens. Effectiveness and mechanisms of oral tolerance in EAU have mainly been studied in the acute, monophasic model in Lewis rats by feeding antigen prior to induction of disease. In this study we investigated the effect of oral tolerance induction in the acute as well as the chronic-relapsing models in the B10.A mouse. In acute murine EAU we could effectively suppress disease by induction of oral tolerance prior to immunization. In the chronic-relapsing EAU, antigen feeding was started only after the animals had recovered from their first attack of uveitis. Under these experimental conditions the subsequent relapse was largely prevented. These experiments demonstrate that oral tolerance may have practical clinical implications in uveitis, which is predominantly a chronic-relapsing condition in humans.
Antibodies to β2-glycoprotein I—a specific marker for the antiphospholipid syndrome
Authors: GUERIN J , FEIGHERY C , SIM R B , JACKSON J
Published: November 2003

The antiphospholipid syndrome is a disorder characterized by recurrent thrombosis and the presence of antibodies specific to phospholipids. However, the diagnosis of this syndrome is hampered by the lack of a specific laboratory test. In this study an ELISA for the measurement of antibodies to solid-phase β2-glycoprotein I (β2-GPI) was established and compared with anticardiolipin antibodies for diagnosis of antiphospholipid syndrome. Significantly elevated levels of antibodies to β2-GPI were found in all patients with definite antiphospholipid syndrome (median  =  91 AU). Marginally elevated levels of antibodies to β2-GPI were observed in 5% of patients with systemic lupus erythematosus (SLE; median  =  4 AU), 1% with stroke (median  =  3 AU), 13% with infectious mononucleosis (median  =  3 AU), 10% with HIV infection (median  =  3 AU) and 8% with VDRL false-positive serology for syphilis (median  =  4 AU), but not in patients with rheumatoid factor, syphilis or carotid artery stenosis. In contrast, significantly raised levels of anticardiolipin antibodies were observed in 100% of patients with definite antiphospholipid syndrome, 30% with SLE, 88% with HIV infection, 94% with syphilis, 62% with infectious mononucleosis, 9% with rheumatoid factor-positive sera, 74% VDRL false-positive serology for syphilis, 47% with stroke and 0% with carotid artery stenosis. This solid-phase assay for antibodies to β2-GPI is highly specific for the antiphospholipid syndrome and represents an advance in the laboratory diagnosis of this disorder.
Regulation of cytokine gene expression by adjuvants in vivo
Authors: VICTORATOS P , YIANGOU M , AVRAMIDIS N , HADJIPETROU L
Published: November 2003

Antibody isotype affects biological activity of the antibodies and therefore should be considered in prevention of disease by vaccination. In previous reports, we demonstrated that adjuvants affect the antibody isotype switching process and favour the production of certain isotypes. The present study extends these findings and shows fundamental differences in the cytokine induction pattern according to the adjuvant used. Cytokine mRNA levels were determined by in situ RNA–RNA hybridization performed on splenocytes isolated from mice injected with different adjuvants. The results revealed that Freund's complete adjuvant (FCA), Freund's incomplete adjuvant (FIA), Al(OH)3 and QuilA administration results in a type-2 (humoral) response, increasing IL-4, IL-5 and IL-13 gene expression, while poly I:C exhibits a type-1 (cell-mediated) response, increasing the production of interferon-gamma (IFN-γ), IL-2 and IL-6 mRNA. Finally, BeSO4 and poly A:U augment IL-5 and IL-6 mRNA production, while lipopolysaccharide (LPS) and LiCl augment IL-6 and tumour necrosis factor-alpha (TNF-α) mRNA production. Also, the adjuvants appear capable of overcoming the inherent IL-2/IFN-γ and IL-4 dichotomy of C57Bl/6 and BALB/c mice, respectively, in response to cellular antigens such as Leishmania and herpessimplex virus (HSV). The overall data suggest that adjuvants direct the isotype switching process via induction of certain cytokines, a finding that can be useful in selection of the most efficient isotype of protective antibodies for disease prevention by vaccination.
IL-13 over-expression in skin is not confined to IgE-mediated skin inflammation
Authors: VAN DER PLOEG I , JEDDI TEHRANI M , MATUSEVICIENE G , WAHLGREN C F , FRANSSON J , SCHEYNIUS A
Published: November 2003

IL-13 is produced by T cells and, like IL-4, it can induce the production of IgE and IgG4. In order to investigate if IL-13 is a specific marker for atopic dermatitis (AD), IL-13 gene expression was analysed in chronic lichenified lesions and non-lesional skin of patients with AD, in involved and non-involved skin of patients with psoriasis, in positive tuberculin reactions in non-atopics, and in the skin of healthy control subjects. Patients with AD (n = 9) showed sensitization to common air-borne allergens (positive Phadiatop) and had total serum IgE values in the range from 10 to 4800 kU/l (median 170 kU/l ). Competitive reverse transcription-polymerase chain reaction (RT-PCR) was used to assess IL-13 gene expression in skin biopsy specimens. IL-13 gene expression was markedly higher in chronic lichenified lesions of patients with AD (P < 0.01), and in the positive tuberculin reactions (P < 0.01; n = 12) than in skin from healthy control subjects (n = 10). However, there was no significant difference in IL-13 gene expression in the skin of patients with psoriasis (n = 10) and that of healthy control subjects. The dermal cell infiltrates were larger and the relative amount of CD3+ and CD4+ cells in these infiltrates was higher in the skin of subjects with a positive tuberculin reaction than in lichenified AD skin. However, these differences were not reflected in differences in IL-13 gene expression. Different triggers of IL-13 gene expression may influence the diverse patterns of inflammation seen in different inflammatory skin disorders.
The production of IL-8 in cerebrospinal fluid in aseptic meningitis of children
Authors: ISHIGURO A , SUZUKI Y , INABA Y , FUKUSHIMA K , KOMIYAMA A , KOEFFLER H P , SHIMBO T
Published: November 2003

Neutrophils accumulate initially in the cerebrospinal fluid (CSF) of aseptic meningitis, perhaps because of increased levels of granulocyte colony-stimulating factor (G-CSF), macrophage inflammatory protein-1α (MIP-1α), and IL-8 in the subarachnoid space. We studied levels of these cytokines in children with aseptic meningitis using ELISA. When meningeal symptoms existed, IL-8 levels (1399 ± 1600 ng/l, n = 32) in the CSF were significantly higher than those either after meningeal symptoms disappeared (61 ± 56 ng/l, n = 18) or in controls (44 ± 63 ng/l, n = 27) ( P < 0.0001). High levels of IL-8 on admission dropped sequentially. Significant correlations were found between IL-8 levels and either neutrophil counts (r = 0.612), G-CSF levels (r = 0.873) or MIP-1α levels (r = 0.623) in the CSF of the affected patients (P < 0.0001). IL-8 values in serum were lower than in the corresponding CSF samples from all individuals with meningeal symptoms. The IL-8 mRNA was detectable by reverse-transcribed polymerase chain reaction (PCR)-assisted amplification in fresh leucocytes from the CSF, but not from the peripheral blood of a healthy volunteer. The culture of CSF mononuclear cells produced high levels of IL-8 (∼ 2750 ng/l). These data indicate that IL-8 levels rise transiently at the initial stage of aseptic meningitis, and that mononuclear cells that migrate into the CSF are a cellular source of this chemokine. We suppose that IL-8, in addition to G-CSF and MIP-1α, contribute to the localized neutrophil accumulation during the disease.
CD28 costimulation and T lymphocyte proliferative responses in HIV-1 infection
Authors: CARLESIMO M , PONTESILLI O , VARANI A R , BERNARDI M L , MAZZONE A M , ROSSO R , GUERRA E C , CASSONE A , PAGANELLI R , AIUTI F
Published: November 2003

To investigate whether defective costimulatory signals could be involved in the loss of T lymphocyte functions during HIV-1 infection, we tested the effect of CD28 costimulation on both T cell receptor/CD3 and HIV-1 antigen-induced proliferative responses. Although CD3-mediated responses significantly decreased with more advanced stages of HIV-1 infection, the ability of potentiating the responses through CD28 costimulation was maintained at all stages and did not differ from that of HIV-1− subjects. When CD28 costimulation was studied in lymphocyte cultures stimulated with HIV-1 gp160 or p24, potentiation was seen only when a significant response was present without additional CD28 triggering, namely in subjects receiving active immunization with recombinant gp160. These results confirm the integrity of the CD28 pathway of costimulation during HIV-1 infection, and suggest that lymphocytes responding to soluble HIV-1 antigen are not deleted in HIV-1-infected patients, but do not receive significant priming during the natural course of the infection.
Induction of human IgE synthesis in B cells by a basophilic cell line, KU812
Authors: YANAGIHARA Y , KAJIWARA K , BASAKI Y , IKIZAWA K , AKIYAMA K , SAITO H
Published: November 2003

Induction of human IgE synthesis in B cells requires, in addition to IL-4 or IL-13, a second signal provided by CD40 ligand (CD40L) on activated Th2-type CD4+ T cells that do not or weakly express Fas ligand (FasL). Mast cells and basophils also produce IL-4 or IL-13 and express CD40L after immunologic or pharmacologic stimulation, although it is unknown whether these cells express FasL. This study investigated the capacity of KU812 cells, a human basophilic cell line, to produce IL-4 and IL-13, to express CD40L and FasL, and to induce IgE and IgG4 synthesis in human normal B cells. Upon stimulation of KU812 cells with either phorbol myristate acetate (PMA) or ionomycin (Iono), IL-4, but not IL-13, was produced in response to Iono, while IL-13, but not IL-4, was inducible by PMA. Moreover, both the time courses of IL-4 and IL-13 production and their amounts secreted were different; IL-4 production was transient, IL-13 production gradually increased, and IL-13 was heavily secreted as compared with IL-4. The combination of PMA and Iono (PMA/Iono) induced higher production of IL-4 or IL-13 than did Iono or PMA alone. KU812 cell-derived IL-4 and IL-13 had the ability to cause CD23 expression on B cells. PMA/Iono also up-regulated CD40L expression and induced a very low level expression of FasL. KU812 cells that had been activated by PMA/Iono followed by fixation could induce IgE and IgG4 synthesis in B cells in the presence of recombinant IL-4 or IL-13. This contact-dependent induction of IgE was completely abrogated by adding anti-CD40L MoAb or soluble CD40, whereas anti-FasL antibody did not significantly affect IgE production. These results indicate that activated KU812 cells produce biologically active IL-4 and IL-13, express functional CD40L, and exhibit weak induction of FasL, thereby supporting sufficient IgE production by B cells.
Differences in the rate of donor leucocyte migration between natural and drug-assisted tolerance following rat liver transplantation
Authors: LORD R , GOTO S , VARI F , EDWARDS-SMITH C , CHIBA S , SUNAGAWA M , LYNCH S V
Published: November 2003

The number involved in and the rate of migration of donor leucocytes into the following recipient organs (spleen, thymus, bone marrow, lung and mesenteric lymph nodes) were measured in two rat models of orthotopic liver transplantation (OLT) using donor-specific MHC class I antibodies. The first OLT model is one that does not require immunosuppression in order to achieve tolerance and involved the transplantation of DA (MHC haplotype, RT1a) livers into PVG (RT1c) recipients. The second model was one that required a 7-day (10 mg/kg) treatment with cyclosporin A (CsA) to achieve tolerance and used DA donors into Lewis (RT11) recipients. Recipient organs were biopsied on days 3, 20 and 87 following OLT and donor leucocyte migration was quantified by immunohistochemistry and computer densitometry of immunoblots of detergent-solublized tissues in order to resolve both membrane-bound and soluble donor MHC class I antigen. In a separate experiment, spleen biopsies were taken following OLT on days 3 and 15 from the naturally tolerizing OLT model (DA into PVG), treated with and without CsA for 7 days and compared with the (DA into Lewis) model. The initial rate of leucocyte migration between days 3 and 21 following OLT was found to be the most rapid into the spleen, followed by the bone marrow and mesenteric lymph nodes in the naturally tolerant (DA into PVG) model when compared with the (DA into Lewis) model. The number of donor leucocytes in the spleen and mesenteric lymph nodes in both models was, however, approximately the same by 87 days. No real difference in the rate of leucocyte migration was seen in the thymus or the lung for both transplant models over the time course assayed. CsA was found to lower the rate of donor leucocyte migration only over the period it was administered. The rate of donor leucocyte migration into the spleen was still much lower 15 days after OLT in the (DA into Lewis) model compared with the (DA into PVG) model treated with and without CsA. Thus the differences in the rate of donor leucocyte migration into the spleen, bone marrow and mesenteric lymph nodes immediately following OLT may offer an explanation as to why the (DA into PVG) combination is able to accept a transplanted liver without immunosuppressive therapy.
High serum level of soluble CD30 in acute primary HIV-1 infection
Authors: PIZZOLO G , VINANTE F , NADALI G , KRAMPERA M , MOROSATO L , CHILOSI M , RAITERI R , SINICCO A
Published: November 2003

CD30 has been suggested to play a role in HIV infection. In this study the serum concentration of soluble CD30 (sCD30) was determined by an ELISA essay on samples collected from patients with acute primary HIV-1 infection during the acute phase (n = 17) and after seroconversion (n = 13). sCD30 during acute infection was consistently elevated (137·58 ± 120·33 versus 6·4 ± 5·4 U/ml (mean ± s.d.) in normal controls; P < 0·0001) and decreased after seroconversion (49·1 ± 66·17 U/ml; P = 0·0018 compared with acute infection). This trend mirrored the disappearance of detectable levels of HIV antigen in the blood, resulting in a direct correlation between sCD30 and HIVAg values (P = 0·002). These data suggest that the high levels of sCD30 observed during the peak concentration of HIVAg in acute primary HIV infection might reflect the high rate of viral replication.
T helper responsiveness in human Loa loa infection; defective specific proliferation and cytokine production by CD4+ T cells from microfilaraemic subjects compared with amicrofilaraemics
Authors: BAIZE S , WAHL G , SOBOSLAY P T , EGWANG T G , GEORGES A J
Published: November 2003

The proliferation and cytokine profiles of peripheral blood mononuclear cells (PBMC) from microfilaraemic (Mf+) subjects infected by Loa loa in response to antigens of several parasitic stages were compared with those from amicrofilaraemic (Mf−) individuals. While a strong lymphoproliferative response and consistent levels of both Th1 (IL-2, interferon-gamma (IFN-γ)) and Th2 (IL-4, IL-5) type cytokines were observed in response to adult worm (AW) and microfilariae (Mf) antigen in Mf− individuals, Mf+ subjects were characterized by a T cell unresponsiveness, including proliferation, cytokine production and IL-2 mRNA expression. Conversely, T cell responsiveness to mitogens and non-specific antigen were similar in the two endemic populations. Depletion of lymphocyte subpopulations indicated that T CD4+ were mainly involved in the specific cellular response. In contrast to other cytokines, IL-10 was produced in response to all parasitic stages, in both Mf+ and Mf− patients. Neutralization of IL-10 did not restore cytokine production in Mf+ patients, while B7 mRNA expression was similar between Mf+ and Mf− subjects in response to Mf antigen, suggesting that IL-10 was not the only factor responsible for T cell unresponsiveness. Mf+ patients have lower Mf antigen-specific IgG levels compared with Mf−, and there is a significant correlation between Mf antigen-specific antibodies and IL-5 responses. These findings suggest that Mf− status is correlated with T helper responsiveness, including proliferation and production of both Th1- and Th2-type cytokines, whereas Mf+ status is characterized by unresponsiveness of the same cell population, induced and/or maintained by microfilariae.
Oxidized low-density lipoprotein (Ox-LDL) but not LDL aggravates the manifestations of experimental antiphospholipid syndrome (APS)
Authors: GEORGE J , BLANK M , HOJNIK M , BAR-MEIR E , KOIKE T , MATSUURA E , LORBER M , AVIRAM M , SHOENFELD Y
Published: November 2003

Ox-LDL is thought to play a major role in atherogenesis. The mechanisms mediating the deleterious influences of Ox-LDL include foam cell formation and cell cytotoxicity. The production of anti-Ox-LDL antibodies results in the formation of immune complexes which are taken up at enhanced rate by macrophages, leading to foam cell formation. APS is characterized by repeated venous and arterial thromboembolic phenomena, recurrent fetal loss and thrombocytopenia, associated with the presence of antibodies to negatively charged phospholipids (aPL) (i.e. cardiolipin, phosphatidylserine). Phospholipids bear structural resemblance to LDL, and several studies have indeed proved that aPL display cross-reactivity with anti-Ox-LDL antibodies. In this study we assessed the capacity of oxidized and native forms of LDL to aggravate the clinical picture of experimentally induced APS in naive mice. Mice were actively immunized intradermally with anticardiolipin antibodies and developed a clinical picture resembling APS in humans. Subsequently, the mice were infused with either Ox-LDL, native LDL or PBS, and similar regimens were applied to controls. APS mice infused with Ox-LDL were found to exhibit a significantly more severe form of the disease in comparison with native LDL- and PBS-infused mice, expressed by lower platelet counts (261 000/mm3, 535 000/mm3 and 455 000/mm3, respectively), longer activated partial thromboplastin time (aPTT) (99 ± 12 s, 63 ± 8 s and 74 ± 8 s, respectively) and higher fetal resorption rates (72·7%, 34·4% and 32·6%, respectively). The results of this study show that Ox-LDL, compared with native LDL, aggravates the clinical manifestations of experimental APS and suggest that cross-reactivity of Ox-LDL with phospholipids may provide a pathogenic explanation for this effect.
Calcium channel blockers suppress the contact hypersensitivity reaction (CHR) by inhibiting antigen transport and presentation by epidermal Langerhans cells in mice
Authors: KATOH N , HIRANO S , KISHIMOTO S , YASUNO H
Published: November 2003

Since Langerhans cells (LC) are the principal antigen-presenting cells among epidermal cells, treatments suppressing LC function may inhibit CHR. Although calcium channel blockers (CCB) have been shown to suppress the functions of several immunologically active cells, little is known about their effect on LC. In this study we show that pretreatment with topical 1% nifedipine or verapamil HCl significantly suppressed both the sensitization and elicitation phases of a CHR in mice. We then investigated whether CCB affected LC. Flow cytometric analysis of regional lymph node cells obtained 24 h after applying FITC demonstrated that topical CCB treatment significantly reduced the percentage of FITC+ NLDC-145+ cells, suggesting that CCB had suppressed antigen transport by LC. In vitro treatment with nifedipine or verapamil significantly suppressed the antigen-presenting capacity of LC in a dose-dependent manner. In addition, in vitro CCB treatment reduced the percentage of class II MHC antigen-positive epidermal cells and significantly suppressed class II MHC and B7-1 levels in LC, as determined by flow cytometry and reverse transcriptase-polymerase chain reaction, whereas surface expression of B7-2 and mRNA was only weakly reduced. Neither expression of CD45 nor the percentage of CD45+ cells were affected, suggesting that the effects of CCB on LC were not due to cytotoxicity. Our results suggest that CCB inhibit CHR, at least in part, by suppressing the functions of LC.
Human blood dendritic cells: binding to vascular endothelium and expression of adhesion molecules
Authors: BROWN K A , BEDFORD P , MACEY M , MCCARTHY D A , LEROY F , VORA A J , STAGG A J , DUMONDE D C , KNIGHT S C
Published: November 2003

To investigate the binding properties of dendritic cells (DC) to vascular endothelium, a comparative analysis was undertaken of DC, monocytes and lymphocytes isolated from the blood of 25 healthy subjects using monolayers of human umbilical vein endothelial cells as the adherence substrate. More blood DC (mean 24% adherence) were adherent to endothelial monolayers than monocytes (mean 18%; P < 0.001) and lymphocytes (mean 12%; P < 0.001). When the monolayers were pretreated with tumour necrosis factor-alpha (TNF-α) all leucocyte populations exhibited an increased attachment, but there was still greater binding of DC (mean 37% adherence) in comparison with monocytes (mean 23%; P < 0.001) and lymphocytes (mean 18%; P < 0.001). Flow cytometric analysis revealed that in relation to monocytes and lymphocytes the DC had a higher surface expression of the adhesion molecules CD11a (P < 0.05), CD11c (P < 0.05) and CD54 (P < 0.05) but a lower prevalence of cells bearing CD49d (mean 38%; P < 0.05) and the homing receptor CD62L (mean 14%; P < 0.001). CD1a was present on 22% of DC and virtually absent from the surface of monocytes and lymphocytes. The intensity of expression of the β1-integrins, CD49c, CD49d and CD49e was greater on DC than lymphocytes and monocytes (P < 0.05). Antibody blocking studies demonstrated that DC binding to untreated and TNF-α-treated endothelium was dependent upon the expression of CD11a, CD18 and CD49d, and the simultaneous application of anti-CD18 and anti-CD49d antibodies produced an approximate 70% inhibition of adhesion (P < 0.001). Thus, the expression of both β1- and β2-integrins contributes to the adhesive interaction between DC and endothelium.
Up-regulation of the granulocyte adhesion molecule Mac-1 by autoantibodies in autoimmune vasculitis
Authors: Johnson P A , Alexander H D , McMillan S A , Maxwell A P
Published: November 2003

The characteristic finding of autoantibodies in patients with vasculitis has raised the possibility that these antibodies play a role in the pathogenesis of the disease. The expression of adhesion molecules (AM) on leucocytes and endothelial cells is believed to be integral to the development of vasculitis. We therefore investigated the effect of sera, positive for anti-neutrophil cytoplasmic antibodies (ANCA) or anti-nuclear antibodies (ANA) from patients with vasculitis, on granulocyte expression of the adhesion molecule Mac-1 (CD11b). Autoantibody-positive sera from 15 out of 35 patients with vasculitis stimulated an up-regulation of Mac-1 on granulocytes. In most cases this effect was reproduced by the autoantibody-positive purified IgG fraction. Autoantibody-negative samples did not stimulate AM up-regulation. Of interest, preincubation of sera with purified antigens did not inhibit AM up-regulation by the autoantibody samples. Blocking the Fc receptors on granulocytes did result in a decrease of Mac-1 up-regulation, but this trend was not statistically significant. These results suggest that both ANCA and ANA have the capacity to up-regulate granulocyte AM expression, and that while Fc interaction with granulocyte Fc receptors is important, it is not the only mechanism whereby such autoantibodies activate cells.
VH genes expressed in peripheral blood IgE-producing B cells from patients with atopic dermatitis
Authors: TILGNER J , GOLEMBOWSKI S , KERSTEN B , STERRY W , JAHN S
Published: November 2003

In this study we analysed the VH genes expressed in IgE-producing peripheral B cells in two patients with severe atopic dermatitis. We found a diverse antibody repertoire containing VH genes of five different VH families, no VH2 and 7 genes, with no biased usage of one particular VH gene family. Some VH germ-line segments were found to be used several times within the ε transcripts (V3–23, V3–30). Comparison of the IgE-encoding VH genes with the respective germ-line elements revealed the expression of nearly germ-line identical as well as somatically mutated VH genes. In these latter VH genes base substitutions were distributed over both complementarity determining and framework regions. Because of the similarities between the IgE committed B cell repertoire in atopic patients and the peripheral B cell repertoire in non-atopic individuals we conclude that the atopic VH gene repertoire probably reflects the result of an increased class switch of polyclonal B cells, including virgin and mature B cells, in response to a Th2 cytokine profile.
A distinct pattern of cytokine production from blood mononuclear cells in multitransfused patients with β-thalassaemia
Authors: SALSAA B , ZOUMBOS N C
Published: November 2003

The unstimulated and induced production of granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), IL-3, IL-6, stem cell factor (SCF), IL-1β, tumour necrosis factor-alpha (TNF-α), TNF-β, interferon-gamma (IFN-γ) and transforming growth factor-beta (TGF-β) was determined after culture of blood mononuclear cells from 22 patients with severe β-thalassaemia in a regular transfusion programme, five non-regularly transfused patients with β-thalassaemia intermedia and nine normal persons. A distinct pattern of cytokine production in thalassaemic patients was detected, namely a low unstimulated production of all cytokines and a significant increase in the stimulated production of IFN-γ, TNF-α and IL-1β; these abnormalities were more pronounced in the more heavily transfused older patients. The increased production of the above cytokines, which usually characterize the acute response to infectious agents and have a negative effect on erythropoiesis, may explain the deterioration of anaemia found in thalassaemic patients during acute infections.
CD44 and hyaluronan expression in the development of experimental crescentic glomerulonephritis
Authors: JUN Z , HILL P A , LAN H Y , FOTI R , MU W , ATKINS R C , NIKOLIC-PATERSON D J
Published: November 2003

CD44 is a widely expressed cell surface glycoprotein which is involved in both cell–matrix and cell–cell interactions which regulate a variety of processes, including leucocyte migration and activation. Therefore, we examined the expression of CD44, and its major ligand hyaluronan, during the induction and progression of experimental glomerulonephritis. Antibody staining of normal rat kidney showed constitutive CD44 expression by resident glomerular macrophages, parietal epithelial cells, medullary and occasional cortical tubules. There was a marked increase in CD44 expression over days 1, 7 and 21 of rat crescentic anti-glomerular basement membrane (GBM) glomerulonephritis. Infiltrating monocytes and lymphocytes were CD44+, with ultrastructural studies showing high levels of CD44 expressed on the surface of lymphocytes adherent to activated endothelium. Marked hyaluronan deposition was seen in areas of fibrosis on days 7 and 21, such as glomerular crescents and the periglomerular area. Hyaluronan deposition was accompanied by the presence of many CD44+ cells. Double immunohistochemistry showed that both CD44+ED1+ macrophages and CD44+ myofibroblasts (identified by expression of α-smooth muscle actin) were present in areas of fibrosis. There was also a dramatic increase in cortical tubular CD44 expression, which was most evident in areas of tubular damage. Although tubular epithelial cells expressed CD44 upon both the basolateral and luminal surface, CD44 expression was most prominent within tight junctions, suggesting a role for CD44–CD44 interactions in cell–cell adhesion within the tubule. Analysis of CD44 isoforms by reverse transcriptase-polymerase chain reaction (RT-PCR) showed that the standard form of CD44 predominated in both normal and diseased kidney. However, a series of alternatively spliced CD44 isoforms was also detected, whose expression was markedly increased during disease. At least seven isoforms containing the v6 domain were identified, with the smallest form representing activated T cells. In conclusion, CD44 is constitutively expressed in normal kidney and is dramatically up-regulated in rat anti-GBM disease, suggesting possible roles for the CD44–hyaluronan interaction in leucocyte recruitment, renal fibrosis and tubular cell–matrix and cell–cell interactions during the induction and progression of crescentic glomerulonephritis.
Further study on the roles of the effector molecules of immunosuppressive macrophages induced by mycobacterial infection in expression of their suppressor function against mitogen-stimulated T cell proliferation
Authors: MAW W W , SHIMIZU T , SATO K , TOMIOKA H
Published: November 2003

Previously, we found that phospholipids and reactive nitrogen intermediates (RNI) collaborated in expression of the T cell mitogenesis-inhibitory activity of immunosuppressive macrophages induced by Mycobacterium avium-intracellulare complex (MAIC) infection. In this study, we examined the roles of free fatty acids (FFA) and prostaglandins (PG) as effectors of MAIC-induced macrophages, and moreover, their collaborating effects with RNI. First, treatment of MAIC-induced macrophages with quinacrine (phospholipase A2 (PLA2) inhibitor), dexamethasone (inhibitor of PLA2 and PG synthesis) or indomethacin (PG synthesis inhibitor) attenuated their suppressor activity against concanavalin A (Con A)-induced mitogenesis of splenocytes (SPC), indicating important roles of FFA liberated from membrane phospholipids and PG, as effectors. Second, oleic acid, PGE2, RNI generated from NOR 4 (a new nitric oxide (NO) donor), and phosphatidylserine (PS) exhibited suppressor activity against SPC mitogenesis without showing significant cytotoxicity, in an irreversible manner. Third, the suppressor activities of RNI and PGE2 were potentiated by combined use with oleic acid in a synergistic manner. Fourth, a dual-chamber experiment in which target SPC were separated from MAIC-induced macrophages by a Millipore filter revealed a requirement for cell-to-cell contact for expression of the suppressor function of MAIC-induced macrophages. These findings indicate that RNI, FFA, PG, and phospholipids (presumably PS) and their collaboration play central roles in expression of the T cell mitogenesis-inhibitory function of MAIC-induced suppressor macrophages.
Lymphocyte activation and subset redistribution in the peripheral blood in acute malaria illness: distinct γδ+ T cell patterns in Plasmodium falciparum and P. vivax infections
Authors: WORKU S , BJÖRKMAN A , TROYE-BLOMBERG M , JEMANEH L , FÄRNERT A , CHRISTENSSON B
Published: November 2003

Lymphocyte subset distributions and activation in the peripheral blood were studied in 39 patients with acute malaria and 16 healthy controls from Addis Ababa and Nazareth, Ethiopia. As confirmed by polymerase chain reaction (PCR), 15 patients were infected with Plasmodium falciparum (Pf ), 17 with P. vivax (Pv) and seven were double-infected (Di) with both Pf and Pv. Three-colour flow cytometry was used for phenotyping. Total leucocyte and lymphocyte counts were lower in malaria patients than in controls. The T cell count was reduced in Pf patients, while in the Pv and Di patients there was a reduction in the natural killer (NK) cell count. The CD4/CD8 ratio remained unchanged. γδ+ T cells were significantly elevated in Pf and Di patients, but not in Pv patients. The increase in γδ+ T cells was mostly due to an increase in Vδ1+ cells. Analyses of cellular activation indicated by the expressions of CD25 and HLA-DR revealed significantly higher numbers of activated CD3+ cells, including γδ+ T cells, in all patient groups compared with controls. Our results thus indicate that in acute malaria illness there is a complex pattern of change in lymphocyte subset distribution and activation, including γδ+ T cells. These patterns in Pf infection seem to be distinct from those in Pv infection.
IgE epitope analysis of the hevein preprotein; a major latex allergen
Authors: BEEZHOLD D H , KOSTYAL D A , SUSSMAN G L
Published: November 2003

We have previously identified the hevein preprotein as a common allergen for latex allergic healthcare workers. The B cell epitopes in the hevein protein that are recognized by IgE of latex-allergic individuals have not been identified. In this study, we examined the hevein preprotein using epitope mapping. Overlapping synthetic peptides of 10 amino acids (two aa overlap) were synthesized on a derivatized cellulose membrane using Fmoc chemistry. The peptide spots were probed with pooled sera from 10 latex-allergic patients, and the IgE-reactive peptides identified with anti-IgE MoAbs. We identified six B cell epitopes within the full length hevein preprotein which bound IgE from latex-allergic patients. Two were located in the N-terminal 5-kD hevein domain and four were observed in the 14-kD C-domain. A broad epitope was located between the N-terminal amino acids 13–24. This epitope had nearly complete homology to wheat germ agglutinin (WGA). Immunological cross-reactivity to WGA was confirmed by Western blot analysis with purified WGA, and this reactivity could be inhibited by latex proteins or WGA. Of the five remaining epitopes, four had homologies to other proteins in the pathogenesis-related family of plant proteins (PR-4). The data demonstrate that hevein has multiple IgE epitopes. The significant homology of these epitopes to a broad family of plant defence proteins further explains the increased prevalence of food allergies in latex-allergic individuals.
Up-regulation of IL-8 secretion by alveolar macrophages from patients with fibrosing alveolitis: a subpopulation analysis
Authors: PANTELIDIS P , SOUTHCOTT A M , BLACK C M , DU BOIS R M
Published: November 2003

Neutrophil accumulation in the lower respiratory tract of patients with fibrosing alveolitis is thought to be facilitated by IL-8, a neutrophil chemoattractant primarily secreted by mononuclear phagocytes. The aims of this study were: (i) to explore IL-8 secretion by lung and blood mononuclear phagocytes in subjects with cryptogenic fibrosing alveolitis, systemic sclerosis with and without fibrosing alveolitis, sarcoidosis and normal individuals; (ii) to examine IL-8 secretory heterogeneity in alveolar macrophages and peripheral blood monocytes; and (iii) to correlate alveolar macrophage phenotypic profile to IL-8 secretion. We observed that more monocytes secreted IL-8 than autologous macrophages and that there was heterogeneity in the in vitro IL-8 secretion by alveolar macrophages and peripheral blood monocytes. IL-8 secretion by alveolar macrophages was significantly higher in subjects with fibrosing alveolitis compared with subjects without fibrosing alveolitis, due to a higher percentage of IL-8-secreting alveolar macrophages in the fibrotic group both in the absence (P<0.002) and presence of lipopolysaccharide (LPS) (P<0.04) and correlated with bronchoalveolar lavage neutrophil percentage. Using the MoAbs RFD1, RFD7 and RFD9, that distinguish subsets of alveolar macrophages, we have been able to identify associations between secretion of IL-8 and smaller cells and the cells identified by the MoAb RFD7. In situ hybridization of the bronchoalveolar lavage cell population revealed that alveolar macrophages are the predominant source of IL-8 in the lung. We conclude that there is an increased number of IL-8-secreting alveolar macrophages in the lungs of patients with fibrosing alveolitis, and IL-8 secretion by these cells is associated with specific phenotypic profile expression.
Coordinate expression of group II phospholipase A2 and the acute-phase proteins haptoglobin (HP) and α1-anti-chymotrypsin (ACH) by HepG2 cells
Authors: VADAS P , GROUIX B , STEFANSKI E , WLOCH M , PRUZANSKI W , SCHROEDER J , GAULDIE J
Published: November 2003

The early response to inflammation is characterized by the synthesis of a variety of proteins under cytokine and glucocorticoid control. During episodes of infection or inflammation, a secretory phospholipase A2 (sPLA2) appears in the circulation along with a variety of acute-phase proteins (APP), suggesting possible common regulatory elements amongst sPLA2 and APP. Using the human hepatoma line, HepG2, regulation of sPLA2 expression was examined in relation to synthesis of HP and ACH. The patterns of induction of sPLA2, HP and ACH were distinct for each of IL-1, tumour necrosis factor (TNF) and IL-6, oncostatin M, IL-11 and leukaemia inhibitory factor. Dexamethasone had an enhancing effect on IL-6-induced expression of HP and ACH, but inhibited sPLA2 expression by 50%. Both 8-bromo-cAMP and dibutyryl cAMP increased sPLA2 expression (48.8-fold and 64.2-fold, respectively), whereas KT5720, an inhibitor of protein kinase A, down-regulated cytokine-induced sPLA2 synthesis by 51%. These data show that a panel of cytokines induced varying patterns of up-regulation of sPLA2, ACH and HP. Although dexamethasone potentiated IL-6-induced APP expression in HepG2 cells, it suppressed sPLA2 expression in a dose-dependent manner. In several respects, sPLA2 regulation is similar to that of HP and ACH, but a notable difference is the reciprocal effect of glucocorticoids on sPLA2 expression compared with that of ACH and HP.
Binding and inhibition of myeloperoxidase (MPO): a major function of ceruloplasmin?
Authors: SEGELMARK M , PERSSON B , HELLMARK T , WIESLANDER J
Published: November 2003

Interactions between plasma proteins and MPO were studied. The protein fraction of normal plasma and serum was shown to exhibit an inhibitory effect on the peroxidase activity of MPO. Most of the inhibitory effect could be retained on an MPO-coupled affinity chromatography column. In particular, a protein with apparent mol. wt of 130 kD showed affinity for MPO. The protein was identified as ceruloplasmin by N-terminal amino acid sequencing and immunochemistry. During separation procedures the peroxidase inhibitory effect was limited to ceruloplasmin-containing fractions of plasma. Purified ceruloplasmin inhibited the peroxidase activity of MPO in a concentration-dependent manner, and exhibited selective binding to MPO-coated microtitre plates. This binding could be inhibited by MPO dissolved in buffer. Correspondingly the binding of MPO to ceruloplasmin-coated plates could be blocked by ceruloplasmin in solution, showing a physical interaction to occur between the two proteins under physiological conditions. We also found affinity to exist between MPO and C3 (and its C3d-containing fragments). However, C3 and C3 fragments did not inhibit the peroxidase reaction in vitro. We propose that ceruloplasmin takes part in the clearance and inactivation of MPO, in vivo. We also speculate that impaired inactivation of MPO may have a pathophysiological role in inflammatory diseases characterized by autoantibodies to MPO, such as rapidly progressive glomerulonephritis with P-ANCA (perinuclear anti-neutrophil cytoplasmic antibodies).
The influence of allotypes on the IgG subclass response to chromosomal β-lactamase of Pseudomonas aeruginosa in cystic fibrosis patients
Authors: CIOFU O , PRESSLER T , PANDEY J P , HØIBY N
Published: November 2003

Sera from 70 adult cystic fibrosis (CF) patients with chronic lung infection with Pseudomonas aeruginosa were typed for seven GM and two KM allotype determinants. IgG class and all four IgG subclasses of antibodies against chromosomal β-lactamase of Ps. aeruginosa (aβab) were measured in all 70 CF patients in a cross-sectional study. The aβab IgG subclass response in sera collected during the first 11 years of chronic infection from 20 CF patients (10 patients with G3M*5 G1M*3/G3M*5 G1M*3 genotype and 10 patients with G3M*21 G1M*1/G3M*21 G1M*1 genotype) was analysed in a longitudinal study. Increased levels of IgG2 were associated with the presence of GM 23 allotype. IgG3 aβab levels were the lowest for subjects with the GM 1,2,3,17 23 5,21 and GM 1,3,17 21 phenotypes and the highest in subjects with GM 3,23,5 and GM 3,5. No significant differences in IgG1 and IgG4 aβab levels were found between the different phenotypes. IgG1 aβab levels were higher in patients with KM*3/KM*3 genotype compared with patients with KM*3, *1 genotype. Patients with G3M*5 G1M*3/G3M*5 G1M*3 genotype had in both the cross-sectional and the longitudinal study higher IgG3 aβab, lower IgG4 aβab levels and poorer lung function than patients with G3M*21 G1M*1/G3M*21 G1M*1 genotype. An influence of the allotypes on the clinical course of chronic lung infection with Ps. aeruginosa in patients with CF is suggested.
Measles virus induces apoptotic cell death in lymphocytes activated with phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore
Authors: ITO M , WATANABE M , IHARA T , KAMIYA H , SAKURAI M
Published: November 2003

Peripheral blood mononuclear cells (PBMC) and T lymphocytes were infected with measles virus (MV) and cultured with a protein kinase C (PKC) activator, PMA and a calcium ionophore, ionomycin. After stimulation, cell viability and incorporation of 5-bromo-2′-deoxyuridine (BrdU) were decreased in MV-infected cells compared with mock-infected cells. DNA content analysis and terminal deoxytransferase (TdT)-mediated dUTP nick end labelling demonstrated that the hypodiploid fraction and DNA fragmentation were increased in MV-infected, T lymphocytes activated with PMA plus ionomycin. These data suggest that MV induces apoptotic cell death in cells activated by PMA plus ionomycin. In contrast to stimulation with PMA plus ionomycin, mitogenic stimulation with phytohaemagglutinin (PHA) did not induce apoptotic cell death in MV-infected cells, although cell proliferation was suppressed. Apoptosis induced in stimulated, MV-infected cells may be one mechanism of immunosuppression.