bluuebunny/crossref_metadata_2025 huggingface dataset:
Suppression of murine collagen‐induced arthritis by nasal administration of collagen
Authors: MYERS L. K. , SEYER J. M. , STUART J. M. , KANG A. H.
Published: November 2003

DBA/1 mice were administered type II collagen (CII) or collagen peptides intranasally before systemic immunization to determine whether tolerance could be induced and autoimmune arthritis suppressed. Although prior experiments have demonstrated that collagen given intravenously or orally is effective, the respiratory mucosal route offers several theoretical advantages for dosing peptides, in addition to ease of use. Intact CII, CB11 and a synthetic peptide containing the immunodominant T‐cell epitope recognized by H‐2q mice were all effective in reducing the incidence and severity of arthritis and the immune response to CII. Since previous studies have demonstrated the importance of IgG2 antibody subclasses to the induction of collagen‐induced arthritis, total immunoglobulin G (IgG), IgG1, and IgG2a and IgG2b were measured. IgG2 antibody subclasses were significantly downregulated by the treatment regimen, whereas a slight decrease in IgG1 antibodies was noted that was not significant. In an effort to determine the mechanism by which arthritis was attenuated, cervical lymph node and spleen cells from treated mice were cultured separately with CII and supernatants tested for the presence of T‐cell lymphokines. The cells provided a T‐helper 2 (Th2)‐like response to CII, with T cells from lymph nodes secreting interleukin‐4 (IL‐4) and splenocytes secreting both IL‐4 and IL‐10, whereas a Th1‐like response was detected in immunized mice not tolerized with CII. These findings indicate that the downregulation of arthritis that occurs with intranasal administration of CII is associated with Th2‐type lymphokine profile and a decrease in complement‐fixing antibody subclass.
A role for interferon‐γ and transforming growth factor‐β in erythroid cell‐mediated regulation of nitric oxide production in macrophages
Authors: SELEDTSOVA G. V. , SELEDTSOV V. I. , TARABAN V. Y. , SAMARIN D. M. , KOZLOV V. A.
Published: November 2003

Interferon‐γ (IFN‐γ) and transforming growth factor‐β (TGF‐β) are known to be a potent inducer and inhibitor for macrophage (Mo) activation process, respectively. In the present study we established that the nucleated erythroid cells (NEC) separated from the spleens of adult (CBA×C57BL/6)F1 (CBF, H‐2k/H‐2d) mice following phenylhydrazine treatment are potentially capable of inducing nitric oxide (NO) production in thioglycollate broth‐elicited peritoneal macrophages (Mo). The stimulating effect of both NEC and their culture supernatant on NO secretion by Mo was most apparent in the presence of bacterial lipopolysaccharide (LPS) and neutralizing antibodies (Abs) to TGF‐β and was largely reversed by the addition to the culture of neutralizing Abs to IFN‐γ. Collectively these results suggest that NEC, through production of IFN‐γ and TGF‐β, may exert a regulatory influence on development and functionality of cells pertaining to monocyte (Mc)/Mo lineage.
Mutation of recombinant complement component C9 reveals the significance of the N‐terminal region for polymerization
Authors: TAYLOR K. M. , TRIMBY A. R. , CAMPBELL A. K.
Published: November 2003

Complement component C9 binds to C5b‐8 sites on target cells and polymerizes to form the membrane attack complex (MAC). The aim of the work reported here was to discover which region within C9 was responsible for protecting the globular protein against self‐polymerization. Computer prediction modelling highlighted the domain at the N‐terminus of C9, which was then investigated by site‐directed mutagenesis. The mutated proteins were expressed using insect cells infected with baculovirus. Removal of 16, 20 or 23 amino acids at the N‐terminus of C9 resulted in inactivation due to self‐polymerization. In contrast, removal of 4, 8 or 12 amino acids resulted in a C9 that did not polymerize spontaneously, had two to threefold enhanced lytic activity on erythrocytes, and had increased binding to C5b‐8 sites on rat neutrophils. These results suggest that the domain within the first 16 amino acids at the N‐terminus of C9 is crucial in preventing the self‐polymerization of the globular protein. We have also found that C9 contains a motif (27WSEWS31) common to a family of cytokine receptors that is similar to a tryptophan‐rich motif (WEWWR) of the membrane pore formers, thiol‐activated cytolysins. Mutation of this motif in C9 resulted in polymerized protein, consistent with this site keeping the N‐terminus in a protected conformation and preventing premature self‐polymerization.
Induction of the 2B9 antigen/dipeptidyl peptidase IV/CD26 on human natural killer cells by IL‐2, IL‐12 or IL‐15
Authors: YAMABE T. , TAKAKURA K. , SUGIE K. , KITAOKA Y. , TAKEDA S. , OKUBO Y. , TESHIGAWARA K. , YODOI J. , HORI T.
Published: November 2003

Activation of human natural killer (NK) cells involves sequential events including cytokine production and induction of cell surface molecules, resulting in the enhancement of cytolytic activity. To delineate the activation process of NK cells, we generated murine monoclonal antibodies (mAbs) against YT, a human large granular lymphocyte/natural killer (LGL/NK) cell line. Among the mAbs reactive with YT cells, one mAb, termed 2B9, was noted because of the lack of reactivity with most of the human T‐ and B‐cell lines tested. In fresh peripheral blood mononuclear cells (PBMC), however, the majority of cells expressing this antigen (Ag) were T cells but not CD16+ nor CD56+ NK cells. Since YT cells showed an activated phenotype expressing interleukin‐2 (IL‐2) receptor α chain, we examined whether 2B9 Ag could be induced on normal human peripheral blood NK cells by cytokines known to activate NK cells. The 2B9 Ag was induced on NK cells by IL‐2, IL‐12 or IL‐15 while no induction was observed by interferon‐γ (IFN‐γ). Biochemical analysis showed that anti‐2B9 mAb recognized a 115 kDa molecule in YT cells. A cDNA clone encoding the 2B9 Ag was isolated from a cDNA expression library of YT cells and its sequence was identical to CD26 cDNA although it was not of full length. Transient expression of the 2B9 cDNA on COS‐7 cells revealed that this cDNA encodes the antigenic epitope(s) recognized by anti‐2B9 mAb as well as Ta1, an anti‐CD26 mAb. These results showed that the 2B9 Ag is identical to CD26, and demonstrated that CD26 is an activation antigen on CD16+ CD56+ NK cells inducible by IL‐2, IL‐12 or IL‐15.
Immunoglobulin response to intrapulmonary immunization of asthmatics
Authors: SCHUYLER M. , LYONS C. R. , MASTEN B. , BICE D.
Published: November 2003

Atopic asthmatics, compared to non‐atopic individuals, exhibit an increased amount of serum antigen‐specific IgE and IgG4 antibody directed toward many aeroallergens. We tested the hypothesis that this difference between atopics and non‐atopics extends to the response to intrapulmonary deposition of a neoantigen, keyhole limpets haemocyanin (KLH). We immunized nine atopic asthmatics and nine non‐atopic controls with 500 μg KLH instilled into a subsegment of the lingula and examined serum anti‐KLH IgG1, IgG2, IgG3, IgG4, IgA1, IgA2 and IgM and specific antibody production by peripheral blood mononuclear cells for 25 days. We also determined specific antibody in bronchoalveolar lavage fluid (BALF) in both the immunized and a non‐immunized lobe 11 days after immunization. We found specific serum antibody in all immunized subjects with no difference between atopics and normals in the amount or kinetics of anti‐KLH IgG1, IgG2, IgG3, IgA1, IgA2 and IgM. However, the atopics exhibited more anti‐KLH IgG4 than the normal controls. Specific anti‐KLH antibody‐producing cells were detected in peripheral blood in most subjects at day 8 to 12 after immunization with no difference between atopics and normals. Specific IgA1, IgA2, IgG1 and IgM antibodies were detected in BALF from the immunized lobes but not from the non‐immunized lobes of both groups of subjects with no difference between atopics and normals. We conclude that atopic asthmatics respond to intrapulmonary KLH with more serum anti‐KLH IgG4 than normal controls, consistent with a bias toward a Th2 response to intrapulmonary exposure to antigen.
Enhancement of B‐cell translocation gene‐1 expression by prostaglandin E2 in macrophages and the relationship to proliferation
Authors: SUK K. , SIPES D. G. , ERICKSON K. L.
Published: November 2003

Although prostaglandin (PG) E2 is known to suppress various macrophage functions, the molecular mechanisms by which that occurs are largely unknown. To understand better those mechanisms, differential screening of a cDNA library from PGE2‐treated macrophages was performed. Subsequently, the DNA sequence of a differentially expressed cDNA clone was determined and the cDNA was identified as B‐cell translocation gene‐1 (BTG1), a recently cloned antiproliferative gene. A two‐ to threefold increase in macrophage BTG1 expression was observed after PGE2 treatment. PGE1 and platelet‐activating factor,but not leukotrienes B4, and C4, or lipopolysaccharide, also enhanced BTG1 expression. Furthermore, this effect was mimicked by dibutyryl cAMP which indicated the involvement of elevated cAMP in the PGE2‐mediated enhancement of BTG1. Moreover, there was an inverse correlation between BTG1 mRNA expression and macrophage proliferation; however, BTG1 alteration was not associated with macrophage tumoricidal activation. Thus, BTG1 may play a role in PGE2‐mediated inhibition of macrophage proliferation and not activation.
Polyclonal Th1 cells transfer oil‐induced arthritis
Authors: SVELANDER L. , MÜSSENER Å. , ERLANDSSON‐HARRIS H. , KLEINAU S.
Published: November 2003

T‐cells play a critical role in oil‐induced arthritis (OIA) in DA rats. The present study focuses on the involvement of CD4/CD8 T cells in OIA by using adoptive transfer. Mitogen‐activated T cells from DA rats previously injected with incomplete Freund’s adjuvant (IFA) were depleted of CD4+ T cells or CD8+ T cells before transfer to irradiated naive receipients. The results indicate that CD4+ T cells are essential for the induction of passively induced OIA. However, in vitro blocking experiments with monoclonal antibodies (mAb) to the CD4 molecule of the T cells before transfer did not affect the passive OIA. Neither was passive OIA inhibited by treating the CD4+ T cells with mAb to intracellular adhesion molecule‐1 (ICAM‐1) in order to block cell–cell interactions or migration. The arthritogenic CD4+ T cells were sensitive, however, to in vitro treatment with mAb to the interleukin‐2 receptor, which inhibited the disease or delayed the onset of passive OIA in recipients. The arthritogenic CD4+ T cells were also analysed for expression of specific T‐cell receptor (TCR) variable (V)β chains, critical for recognition of autoantigen, by utilizing Vβ gene‐specific polymerase chain reaction (PCR). The results show a heterogeneous expression of Vβ segments of the TCR, indicating a polyclonal origin of the pathogenic cells. Moreover, an investigation of the T helper (Th)1/Th2 status of the CD4+ T cells, defined by cytokine expression, was made at the mRNA level by using in situ hybridization. High numbers of interleukin‐2 (IL‐2) mRNA expressing cells and also interferon‐γ (IFN‐γ) and tumour necrosis factor‐α (TNF‐α)‐expressing cells could be identified. We conclude from this study that non‐immunogenic IFA triggers polyclonal, IL‐2‐dependent Th1 cells which induce arthritis. The contribution of the CD4 or ICAM‐1 molecules for arthritis induction seem to be of minor importance.
Regulation of HLA class I and II expression by interferons and influenza A virus in human peripheral blood mononuclear cells
Authors: KESKINEN P. , RONNI T. , MATIKAINEN S. , LEHTONEN A. , JULKUNEN I.
Published: November 2003

HLA class I and II molecules play a central role in regulating host immune responses against microbial infections because they present foreign antigens to CD8+ and CD4+ T lymphocytes, respectively. Many cytokines, especially interferons (IFN), are known to upregulate human leucocyte antigen (HLA) class I and II gene expression, but the kinetics, expression levels and viral regulation of HLA genes in primary human cells have not been well documented. Stimulation of peripheral blood mononuclear cells (PBMC) with IFN‐α and IFN‐γ resulted in a 1·5‐ to twofold increase in HLA class I and β2‐microglobulin expression in lymphocytes and monocytes. Lymphocytes did not express any detectable HLA class II either basally or after IFN induction. In monocytes, instead, a high basal class II expression was found and it was further induced by IFN‐α (up to twofold) and especially by IFN‐γ (up to fivefold). In granulocyte–macrophage colony‐stimulating factor (GM‐CSF) differentiated human macrophages, basal HLA class I and II protein expression levels were high but IFN‐γ stimulation was able to further enhance their expression. Accordingly, class I and II mRNA expression was elevated by IFN‐γ, whereas IFN‐α practically had no effect on HLA class I mRNA levels. Influenza A virus infection of macrophages resulted in temporary increases in HLA class I, β2‐microglobulin and class II antigen expression. Neutralization of virus‐induced IFN production by antibodies against type I and II IFNs prevented the virus‐induced upregulation of HLA antigens. At late times of infection, as analysed by steady‐state mRNA expression, both HLA class I and II mRNA were strongly reduced. These results suggest that IFNs are important regulators of HLA genes and responsible for a temporary increase in HLA antigen expression during influenza A virus infection.
Interleukin‐13 selectively induces monocyte chemoattractant protein‐1 synthesis and secretion by human endothelial cells. Involvement of IL‐4Rα and Stat6 phosphorylation
Authors: GOEBELER M. , SCHNARR B. , TOKSOY A. , KUNZ M. , BRÖCKER E.‐B. , DUSCHL A. , GILLITZER R.
Published: November 2003

Chemokines secreted by endothelium have been demonstrated to promote leucocyte recruitment to sites of inflammation. In the present study we investigated the effect of the T lymphocyte‐secreted cytokine interleukin (IL)‐13 on endothelial expression of chemokines. Employing in situ hybridization and enzyme‐linked immunosorbent assay (ELISA) techniques we demonstrate that IL‐13, which shares many of its activities with IL‐4, selectively induces expression of the C‐C chemokine monocyte chemoattractant protein (MCP)‐1 in human umbilical vein endothelial cells (HUVEC). However, it fails to up‐regulate other C‐C and C‐X‐C chemokines potentially inducible in endothelium such as RANTES (regulated on activation, normal Texpressed and secreted), gro‐α, or IL‐8. IL‐13 dose‐dependently induces monocyte chemotactic activity by HUVEC which can be efficiently blocked by neutralizing antisera against MCP‐1. In contrast to the synergistic effect of IL‐13 and tumour necrosis factor‐α (TNF‐α) on endothelial vascular cell adhesion molecule‐1 (VCAM‐1) surface expression, TNF‐α‐induced secretion of MCP‐1 is not augmented by IL‐13. Studying the signalling pathway activated by IL‐13 it is demonstrated that a neutralizing monoclonal antibody (mAb) to the 140 000 MW component of the IL‐4 receptor (IL‐4Rα) inhibits the effect of IL‐13. Immunoprecipitation studies reveal that endothelial IL‐4Rα is rapidly tyrosine phosphorylated upon treatment with IL‐13 and IL‐4. We furthermore show that both cytokines activate the signal transducer and activator of transcription (Stat) protein‐6 in endothelial cells. Our data suggest that IL‐13 partly utilizes components of the IL‐4 receptor signalling pathway for induction of endothelial MCP‐1 expression to facilitate recruitment of blood leucocytes.
Differential functional effects of a humanized anti‐CD4 antibody on resting and activated human T cells
Authors: BRETT S. J. , ROWAN W. , SMITH M. , BARTHOLOMEW M. , TITE J. P.
Published: November 2003

A fully humanized immunoglobulin G1 (IgG1) anti‐CD4 monoclonal antibody is currently being evaluated in phase I/II clinical trials for rheumatoid arthritis. In order to understand the mode of action of this antibody in vivo, we have carried out a detailed functional analysis in vitro of the effects of this antibody on T‐cell activation. The anti‐CD4 antibody was found to inhibit both antigen‐specific responses involving recognition of human leucocyte antigen (HLA) class II and processed antigenic peptides as well as non‐class II dependent responses via anti‐CD3 antibodies. The antibody did not cause total blockade of T‐cell proliferation, but rather induced a shift in the dose–response curve, decreasing the sensitivity of cells to antigen or anti‐CD3‐mediated stimulation. The antibody appears to allow at least a partial early signal into the T cell as it does not inhibit the increase in tyrosine phosphorylation induced by anti‐CD3 antibodies. A comparison of the intact antibody with that of either the F(ab′)2 fragment or an engineered non‐Fc receptor (FcR) binding form revealed that the intact antibody was the most effective at inhibiting proliferation of resting peripheral blood CD4+ T cells. However, this difference was only apparent when excess antibody was removed from culture prior to antigen or anti‐CD3 mediated stimulation. The intact antibody induced both CD4 down‐modulation and increases in CD4‐associated tyrosine phosphorylation of resting CD4+ T cells, which were not seen with the non‐FcR binding versions, which may account for the enhanced potency of the intact antibody at inhibiting T‐cell activation. Interestingly, the anti‐CD4 antibody induced a differential effect on activated CD4+ T cell clones compared with resting CD4+ T cells with respect to degree of CD4 cross‐linking required to induce functional effects in the T cell. Both intact and non‐FcR binding antibodies were equally effective at inhibiting T‐cell proliferation of activated T‐cell clones. In addition CD4 down‐modulation and increased CD4‐associated tyrosine phosphorylation were observed with T‐cell clones in the absence of secondary cross‐linking. Such observations may be of relevance when studying the effects of the antibody at sites of inflammation, where there will be CD4+ T cells of differing activation states as well as varying numbers of FcR positive cells.
Enhancement of CD4+ T‐cell‐dependent interleukin‐2 production in vitro by murine alveolar macrophages: the role of leukotriene B4
Authors: MARCINKIEWICZ J. , GRABOWSKA A. , BRYNIARSKI K. , CHAIN B. M.
Published: November 2003

Local tissue macrophages are known to play a key role in regulation of adaptive immune responses, often by inhibition of T‐cell activation and proliferation. In this study, we compare the influence of alveolar and peritoneal macrophages on T‐cell‐dependent interleukin‐2 (IL‐2) release. Alveolar macrophages, in contrast to peritoneal macrophages, enhance IL‐2 release. Assay of a panel of potential macrophage‐derived mediators indicated that activated alveolar macrophages stimulated greater release of IL‐1β, tumour necrosis factor‐α and, especially, leukotriene B4 (>100 times) than activated peritoneal macrophages. Inhibition of prostaglandin synthesis by alveolar macrophages further enhanced the production of IL‐2, while inhibition of leukotriene synthesis abolished the enhancement. The addition of exogenous prostaglandin E2 inhibited IL‐2 release, while exogenous leukotriene B4 enhanced IL‐2 release. When added simultaneously, the two compounds antagonized each other’s activity. In conclusion, this study confirms that alveolar macrophages enhance IL‐2 secretion, and suggests that this enhancement may be due at least in part to the very high rates of production of leukotriene B4. The overall influence of macrophage populations on T cells in vivo will reflect the complex balance between the multiple mediators produced within the local tissue microenvironment.
Changes in swine macrophage phenotype after infection with African swine fever virus: cytokine production and responsiveness to interferon‐γ and lipopolysaccharide
Authors: WHITTALL J. T. D. , PARKHOUSE R. M. E.
Published: November 2003

Cytokines produced by cells of the immune system, including macrophages, can influence inflammatory responses to viral infection. This has been exploited by viruses, which have developed strategies to direct the immune response towards ineffective responses. African swine fever virus (ASFV) is a double‐stranded DNA virus that infects macrophages of domestic swine. In this study, primary cells of monocyte–macrophage lineage were obtained from the lungs, peritoneum or blood of domestic swine and, after infection with ASFV, supernatants were tested for cytokines using biological assays. The cytokine transforming growth factor‐β (TGF‐β) was detected after infection of macrophage preparations, but tumour necrosis factor (TNF) and interleukin‐1 (IL‐1) were not detected. ASFV‐infected and uninfected macrophage populations were also tested to assess their ability to respond to cytokines by enhancing production of superoxide in the respiratory burst mechanism. Responses to interferon‐γ (IFN‐γ) and lipopolysaccharide (LPS) were suppressed in macrophage populations infected with virus, even at low multiplicities of infection. Addition of TGF‐β to uninfected macrophages resulted in a similar suppression of response, but antibody to TGF‐β did not prevent suppression induced by virus. These results are discussed in relation to the pathology of African swine fever.
Immune memory in CD4+ CD45RA+ T cells
Authors: RICHARDS D. , CHAPMAN M. D. , SASAMA J. , LEE T. H. , KEMENY D. M.
Published: November 2003

This study addresses the question of whether human peripheral CD4+ CD45RA+ T cells possess antigen‐specific immune memory. CD4+ CD45RA+ T cells were isolated by a combination of positive and negative selection. Putative CD4+ CD45RA+ cells expressed CD45RA (98·9%) and contained <0·1% CD4+ CD45RO+ and <0·5% CD4+ CD45RA+ CD45RO+ cells. Putative CD45RO+ cells expressed CD45RO (90%) and contained 9% CD45RA+ CD45RO+ and <0·1% CD4+ CD45RA+ cells. The responder frequency of Dermatophagoides pteronyssinus‐stimulated CD4+ CD45RA+ and CD4+ CD45RO+ T cells was determined in two atopic donors and found to be 1 : 11 314 and 1 : 8031 for CD4+ CD45RA+ and 1 : 1463 and 1 : 1408 for CD4+ CD45RO+ T cells. The responder frequencies of CD4+ CD45RA+ and CD4+ CD45RO+ T cells from two non‐atopic, but exposed, donors were 1 : 78 031 and 1 : 176 903 for CD4+ CD45RA+ and 1 : 9136 and 1 : 13 136 for CD4+ CD45RO+ T cells. T cells specific for D. pteronyssinus were cloned at limiting dilution following 10 days of bulk culture with D. pteronyssinus antigen. Sixty‐eight clones were obtained from CD4+ CD45RO+ and 24 from CD4+ CD45RA+ T cells. All clones were CD3+ CD4+ CD45RO+ and proliferated in response to D. pteronyssinus antigens. Of 40 clones tested, none responded to Tubercule bacillus purified protein derivative (PPD). No difference was seen in the pattern of interleukin‐4 (IL‐4) or interferon‐γ (IFN‐γ) producing clones derived from CD4+ CD45RA+ and CD4+ CD45RO+ precursors, although freshly isolated and polyclonally activated CD4+ CD45RA+ T cells produced 20–30‐fold lower levels of IL‐4 and IFN‐γ than their CD4+ CD45RO+ counterparts. Sixty per cent of the clones used the same pool of Vβ genes. These data support the hypothesis that immune memory resides in CD4+ CD45RA+ as well as CD4+ CD45RO+ T cells during the chronic immune response to inhaled antigen.
Superantigenicity of helper T‐cell mitogen (SPM‐2) isolated from culture supernatants of Streptococcus pyogenes
Authors: RIKIISHI H. , OKAMOTO S. , SUGAWARA S. , TAMURA K. , LIU Z‐X. , KUMAGAI K.
Published: November 2003

A superantigen (Streptococcus pyogenes mitogen‐2; SPM‐2) that stimulates human helper T cells bearing unique types of variable domains of T‐cell receptor β‐chain (TCR Vβ) was isolated from the culture supernatant of S. pyogenes strain T12. The active molecule isolated by diethylaminoethyl (DEAE)–cellulose chromatography and isoelectric focusing was a protein with a molecular weight (MW) of 29 000 and isoelectric point (pI) of 6·0. This new superantigen was found to activate preferentially Vβ4+, 7+, and 8+ T cells, whereas recombinant streptococcal pyrogenic exotoxin A and C activated Vβ12+ and Vβ2+ T cells, respectively, as determined by flow cytometry and reverse transcriptase‐polymerase chain reaction (RT–PCR) methods. This proliferative response was significantly inhibited by anti‐HLA‐DR monoclonal antibody, and required paraformaldehyde‐fixed antigen‐presenting cells (APC), indicating that this action is dependent on major histocompatibility complex (MHC) class II molecules without processing. Analysis of the amino‐terminal amino acid sequence of the molecule failed to find any identical or significantly homologous proteins. We have previously reported that cytoplasmic membrane‐associated protein (CAP), a streptococcal superantigen isolated from the cell membranes of S. pyogenes T12 strain, stimulated mainly Vβ8+ T cells. Both SPM‐2 and CAP preferentially stimulated helper T cells, and rabbit antiserum against SPM‐2 completely neutralized the T‐cell‐stimulating activities of CAP, suggesting that SPM‐2 and CAP belong to a family of streptococcal mitogenic proteins. The SPM‐2 activity with stimulation of Vβ8+ T cells was detected extensively in the culture fluids of group A streptococci, but not in those of other streptococcal species, including groups B and D streptococci, and most of the activities detected were completely inhibited by anti‐SPM‐2 serum. These results indicate that SPM‐2 may be a newly discovered superantigen molecule, which can be commonly synthesized by group A streptococci.
Selective enhancement of endothelial cell VCAM‐1 expression by interleukin‐10 in the presence of activated leucocytes
Authors: FIEHN C. , PALEOLOG E. M. , FELDMANN M.
Published: November 2003

Although initially described as an immunomodulatory cytokine, interleukin‐10 (IL‐10) has also been proposed to exert proinflammatory effects both in vivo and in vitro. In particular, studies in IL‐10 transgenic mice have suggested that IL‐10 may activate vascular endothelium to promote leucocyte adhesion and extravasation. In the present study we investigated whether IL‐10 activates endothelial cells either directly or indirectly, via signals produced by leucocytes in the endothelial cell environment, using a co‐culture of human umbilical vein endothelial cells and peripheral blood mononuclear cells (PBMC). No direct effects of IL‐10 on endothelial cell responses were observed. However, in the presence of phytohaemagglutinin‐activated PBMC, IL‐10 increased the expression on endothelial cells of vascular cell adhesion molecule‐1 (VCAM‐1) but not of intercellular adhesion molecule‐1, E‐selectin or major histocompatibility complex (MHC) antigens, an effect mediated by PBMC‐derived soluble factors. We also observed that interferon‐γ (IFN‐γ) antagonized VCAM‐1 expression on endothelial cells mediated by IL‐4 and IL–13. Since IL‐10 has previously been documented to down‐regulate release of IFN‐γ by PBMC, we propose that the IL‐10‐mediated reduction of IFN‐γ production by PBMC results in enhanced responsiveness of endothelial cells to PBMC‐derived IL‐4 and IL‐13, and thus increased expression of VCAM‐1. Our results suggest that the relative balance of cytokines produced by infiltrating cells in developing inflammatory lesions may differentially modulate endothelial responsiveness in vivo, and that IL‐10 might indirectly stabilize VCAM‐1 expression on endothelial cells by affecting the balance of leucocyte‐derived cytokines in the endothelial environment.
Interleukin‐10 modulates susceptibility in experimental cerebral malaria
Authors: KOSSODO S. , MONSO C. , JUILLARD P. , VELU T. , GOLDMAN M. , GRAU G. E.
Published: November 2003

In this study, we examined the effects of interleukin‐10 (IL‐10) on the outcome of experimental cerebral malaria (CM), a lethal neurological syndrome that occurs in susceptible strains of mice after infection with Plasmodium berghei ANKA (PbA). Constitutive IL‐10 mRNA levels were significantly higher in the spleen and brain of resistant animals. In vivo neutralization of endogenous IL‐10 in CM‐resistant mice induced the neurological syndrome in 35·7% of these mice, as opposed to 7·7% in controls. IL‐10 inhibited PbA antigen‐specific interferon‐γ (IFN‐γ) production in vitro but not tumour necrosis factor (TNF) serum levels in vivo. Susceptible mice, on the other hand, were significantly protected against CM when injected with recombinant IL‐10. Overall, our findings suggest that IL‐10 plays a protective role against experimental cerebral malaria.
HIV‐1‐specific cell‐mediated immune responses induced by DNA vaccination were enhanced by mannan‐coated liposomes and inhibited by anti‐interferon‐γ antibody
Authors: TODA S. , ISHII N. , OKADA E. , KUSAKABE K.‐I. , ARAI H. , HAMAJIMA K. , GORAI I. , NISHIOKA K. , OKUDA K.
Published: November 2003

The adjuvant effect of mannan‐coated liposomes on human immunodeficiency virus type‐1 (HIV‐1) DNA vaccine and the mechanism of this enhancement were studied. Coating of cationic liposomes with mannan significantly enhanced the ability of this vaccine to induce an HIV‐specific delayed‐type hypersensitivity (DTH) response. HIV‐specific cytotoxic T‐cell (CTL) activity elicited by DNA vaccination was also significantly enhanced with the mannan‐liposome cocktail. This mannan‐liposome‐mediated activity was greatly inhibited by in vivo injection of anti‐interferon (IFN)‐γ antibody, which suggests that IFN‐γ plays an important role in this HIV‐specific immune response. The results of both isotype‐specific antibody and cytokine analysis revealed that mannan‐liposome‐mediated DNA vaccination enhances Th1‐mediated immunity.
Differential expression of CD8 epitopes amongst porcine CD8‐positive functional lymphocyte subsets
Authors: YANG H. , PARKHOUSE R. M. E.
Published: November 2003

The swine is a useful model for immunobiological studies as it has a highly heterogeneous lymphocyte pool, containing several subsets not easily accessible in humans and rodents. In particular, the CD8‐positive (CD8+) cells contain a variety of lymphocyte subsets, such as αβ‐T cells, γδ‐T cells, CD4 CD8 double‐positive (DP) cells and natural killer (NK) cells. In order to define these subsets further, we have selected four monoclonal antibodies (mAb) with differential reactivity on CD8+ cells. Thus, mAb CD8.1 (PPT20) bound to CD8hi and CD8lo subpopulations in a similar way to the conventional anti‐CD8. The mAb CD8.2 (PPT21), though binding to all of the CD8+ cells, reacted preferably with CD8hi. Two other mAb, CD8.3 (PPT22) and CD8.4 (PPT23), were specific for CD8hiαβ‐T‐cell subpopulation. These results, complemented by immunoprecipitation, co‐modulation and enzyme‐linked immunosorbent assay experiments, suggest that CD8.1 and CD8.2 react putatively with the CD8α‐chain and CD8.3 and CD8.4 with the CD8β‐chain. Tissue distribution studies revealed that CD8+ thymocytes and peripheral CD8hiαβ‐T cells expressed both putative CD8α‐ and β‐chains while peripheral CD4+CD8+αβ‐T cells, CD8loγδ‐T cells and NK cells expressed only putative CD8α‐chain. Functional studies indicated that the CD8hiαβ‐T and CD8loγδ‐T cells were effector cells in the CD3‐redirected cytotoxicity.
Identification of specific recognition molecules on murine mononuclear phagocytes and B lymphocytes for Vi capsular polysaccharide: modulation of MHC class II expression on stimulation with the polysaccharide
Authors: QADRI A.
Published: November 2003

Vi bacterial polysaccharide is a homopolymer of α1–4 N‐acetyl polygalacturonic acid with variable O‐acetylation at position C‐3 and forms a capsule around many bacteria. It has been referred to as the virulence factor of Salmonella typhi and is also a candidate vaccine against typhoid fever. The present study reports the interaction of this polysaccharide with murine mononuclear phagocytes and lymphocytes, and with human monocytes. Vi showed a dose‐dependent binding to the murine monocyte cell lines WEHI‐274.1 and J774. This binding was abrogated if the polysaccharide was deacetylated, suggesting involvement of acetyl groups in this interaction. Vi also bound to the murine B‐cell lymphoma line A20, to peritoneal exudate cells and to a lesser degree to spleen cells and thymocytes from BALB/c mice. The polysaccharide also interacted with the human histiocytic lymphoma line U937 but not with the human monocyte cell line THP‐1. Stimulation with Vi led to up‐regulation of surface major histocompatibility complex (MHC) class II expression on A20 cells. Immunoprecipitation of Vi‐bound molecules from cell surface biotinylated A20 and WEHI‐274.1 revealed two bands with MW of about 32 000 and 36 000. The study demonstrates that Vi capsular polysaccharide can interact with mononuclear phagocytes and lymphocytes through specific cell surface molecules and modulate MHC class II expression.
GM‐CSF increases the ability of cultured macrophages to support autologous CD4+ T‐cell proliferation in response to Dermatophagoides pteronyssinus and PPD antigen
Authors: CAULFIELD J. J. , HAWRYLOWICZ C. M. , KEMENY* D. M. , LEE T. H.
Published: November 2003

Previous studies have demonstrated an infiltration of monocytes and increased levels of granulocyte–macrophage colony‐stimulating factor (GM‐CSF) in the asthmatic lung. To study the possible effects of this cytokine upon the differentiation and function of these newly recruited monocytes, we have developed a model in which monocytes isolated from human peripheral blood were differentiated into macrophages in serum in the presence or absence of GM‐CSF. After 7 days, the macrophages increased in size and granularity, had increased phagocytic activity, and expressed various adhesion molecules, CD14 and major histocompatibility complex (MHC) class II. The effects of GM‐CSF on antigen presentation by cultured macrophages on the antigen‐ specific proliferative response of CD4+ T cells to Dermatophagoides pteronyssinus or purified protein derivative of tuberculin and the mitogen phytohaemagglutinin was determined. CD4+ T‐cell proliferation was reduced when either antigen was presented by macrophages cultured in serum alone, compared with the values obtained with freshly isolated monocytes. However, CD4+ cell proliferation was comparable to that observed with monocytes when antigen was presented by macrophages which had been pre‐cultured with 50 U/ml GM‐CSF. CD4+ T‐cell proliferation to phytohaemagglutinin was similar when all three populations were used as accessory cells. High numbers of macrophages partially suppressed CD4+ T‐cell proliferation in response to antigen presented by monocytes, but there was no significant difference between macrophages cultured in the presence or absence of GM‐CSF. This data suggests that GM‐CSF directs monocyte differentiation into macrophages with an antigen‐presenting, rather than a suppressive, phenotype. Elevated levels of GM‐CSF in the asthmatic lung may therefore maintain recently recruited monocytes in an inflammatory and T‐cell activating state.
The species‐specific structure of microanatomical compartments in the human spleen: strongly sialoadhesin‐positive macrophages occur in the perifollicular zone, but not in the marginal zone
Authors: STEINIGER B. , BARTH P. , HERBST B. , HARTNELL A. , CROCKER P. R.
Published: November 2003

The microanatomical structure of human and rat splenic white pulp is compared, with special emphasis on the localization of the marginal zone occupied by immunoglobulin M (IgM)+ IgD−/dull B lymphocytes and its specialized macrophages. Our study reveals that in contrast to rats, the marginal zone of humans primarily exists in the vicinity of primary and secondary splenic follicles and that it is almost absent around the periarteriolar T‐cell zones. We demonstrate that in humans there is an additional compartment, the perifollicular zone, located between the marginal zone and the red pulp. The perifollicular zone is a dynamic region of variable cellular and phenotypic composition, which can be regarded either as a part of the red pulp or of the follicles. In most cases the perifollicular zone appears as a compartment of the red pulp containing erythrocyte‐filled spaces which differ from the typical red pulp sinusoids. Similar to the splenic cords, the perifollicular zone mostly harbours scattered B and T lymphocytes. However, sometimes B lymphocytes clearly predominate in the perifollicular area. In addition, strongly sialoadhesin‐positive macrophages form sheaths around capillaries in the perifollicular zone. Such capillary sheaths are not observed in rats. In humans weakly sialoadhesin‐positive macrophages are also present in the perifollicular zone and in the red pulp. In some specimens sialoadhesin is, however, strongly expressed by a large number of dispersed perifollicular macrophages. Interestingly, in striking contrast to rats, the human marginal zone does not contain sialoadhesin‐positive macrophages and marginal metallophilic macrophages are also absent in humans. Thus, sialoadhesin‐positive macrophages and IgM+ IgD− memory B lymphocytes both share the marginal zone as a common compartment in rats, while they occupy different compartments in humans. We show that the human splenic marginal zone does not contain a marginal sinus and assume that in humans the perifollicular region is the compartment where antigen and recirculating lymphocytes enter the organ.
Tosylphenylalanine chloromethyl ketone inhibits TNF‐&agr; mRNA synthesis in the presence of activated NF‐&kgr;B in RAW 264.7 macrophages
Authors: JEONG J.‐Y. , KIM K.‐U. , JUE D.‐M.
Published: November 2003

Serine proteinase inhibitors such as N‐tosyl–phenylalanine chloromethyl ketone (TPCK) and N&agr;‐p‐tosyl–lysine chloromethyl ketone (TLCK) were shown to inhibit production of tumour necrosis factor‐&agr; (TNF‐&agr;) in lipopolysaccharide (LPS)‐activated RAW 264.7 macrophages. The proteinase inhibitors were also reported to inhibit activation of the transcription factor nuclear factor‐&kgr;B (NF‐&kgr;B) by blocking the signalling pathway for stimuli‐induced phosphorylation of the inhibitory subunit (I&kgr;B&agr;) and thus preventing its degradation. In RAW 264.7 cells TPCK and TLCK significantly suppressed LPS‐induced increase in TNF‐&agr; mRNA, induction of nuclear &kgr;B‐binding activity and degradation of I&kgr;B&agr;. TPCK and TLCK effectively blocked TNF‐&agr; mRNA synthesis even when they were added after LPS stimulation. In these cells, however, the inhibitory modes of the two inhibitors were found to be different: while addition of TLCK suppressed I&kgr;B&agr; degradation and reduced NF‐&kgr;B activity, a comparable decrease in the nuclear &kgr;B‐binding activity or in I&kgr;B&agr; degradation was not observed in cells treated with TPCK. Our results show that TPCK inhibits LPS‐induced TNF‐&agr; mRNA synthesis in the presence of activated NF‐&kgr;B and suggests that mechanisms other than NF‐&kgr;B activation are involved in the transcriptional regulation of the TNF‐&agr; gene.
Macrophages activated by Listeria monocytogenes induce organ‐specific autoimmunity
Authors: SONODA K.‐H. , MATSUZAKI G. , NOMURA A. , YAMADA H. , HAMANO S. , NAKAMURA T. , MUKASA A. , NOMOTO K.
Published: November 2003

We have previously reported an experimental autoimmune model induced by the local infection of Listeria monocytogenes. The unilateral inoculation of virulent Listeria into a testis of a normal mouse induced a delayed‐type hypersensitivity response against testicular antigen and caused autoimmune orchitis in the contralateral testis. The orchitis was transferred to naive mice by T cells from the intratesticularly infected mice. In this paper, we demonstrated that avirulent Listeria, which lacks the expression of listeriolysin O, failed to induce any anti‐testicular responses or contralateral orchitis even when it was inoculated at a high dose into the testis. Furthermore, the intraperitoneal inoculation of virulent Listeria with testicular antigen induced the anti‐testicular responses and orchitis although intraperitoneal inoculation of testicular antigen with avirulent Listeria failed to induce them. The difference between virulent and avirulent Listeria in the induction of anti‐testicular responses was supposed to be dependent on the difference in macrophage activation by the two bacterial strains because, first, the anti‐testicular responses were elicited in normal mice when macrophages from virulent Listeria‐infected mice were intraperitoneally transferred with testicular antigen although no viable bacteria were detected from the macrophages, and secondly, in contrast, the intraperitoneal co‐inoculation of macrophages from avirulent Listeria‐infected mice and testicular antigen failed to elicit any anti‐testicular responses. Finally, we found that the virulent Listeria‐induced macrophages expressed a higher level of CD80 (B7‐1) and CD86 (B7‐2) molecules than did the avirulent Listeria‐induced macrophages and naive peritoneal macrophages. These results thus suggest that virulent Listeria activates macrophages to induce autoreactive T cells while avirulent Listeria does not. The up‐regulation of B7 molecules by virulent Listeria infection is a candidate of the mechanism for the activation of autoreactive T cells.
Both CD4+ and CD8+ human lymphocytes are activated and proliferate in response to Cryptococcus neoformans
Authors: SYME R. M. , WOOD C. J. , WONG H. , MODY C. H.
Published: November 2003

The current studies were performed to determine the contribution of T‐cell subsets to lymphocyte proliferation in response to Cryptococcus neoformans, the most common invasive mycosis in acquired immune deficiency syndrome. We demonstrate for the first time that both human CD4 and CD8 cells are activated in response to C. neoformans. Both CD4 and CD8 cells express interleukin‐2 receptor &agr; (IL‐2R&agr;) and transferrin receptor and proliferate in response to C. neoformans, however proliferation of CD8 cells was dependent upon CD4 cells. The requirement for CD4 cells was complex, since CD8‐enriched cells failed to express mRNA for IL‐2, suggesting that CD4‐dependent IL‐2 production was required for CD8‐cell proliferation. However, IL‐2 was not sufficient to restore CD8‐cell proliferation. These studies provide experimental evidence in humans to support the clinical impression that CD4 cells are important in cryptococcosis, and suggest that the appropriate CD4‐derived signals could allow CD8 cells to assist in host defence.
Identification of two distinct subsets of bovine γδ T cells with unique cell surface phenotype and tissue distribution
Authors: MACHUGH N. D. , MBURU J. K. , CAROL M. J. , WYATT C. R. , ORDEN J.A. , DAVIS W. C.
Published: November 2003

We describe the characterization of two subsets of bovine γδ T cells having distinct cell surface phenotype and tissue distribution. One population expresses the previously described 215 000 MW WC1 antigen and is negative for the cell‐surface differentiation antigens CD2, CD4, and CD8. The second population expresses CD2 and CD8 but not WC1 and appears to have a T‐cell receptor (TCR) rearrangement distinct from that of the WC1+ population. The WC1− population is found in large numbers in spleen and intestine. In addition, this subset is not recognized by a number of monoclonal antibodies (mAbs) specific for TCR families that are well represented in the WC1+ population. The results indicate that the γδ T‐cell population in cattle is considerably larger than previously described and that this population can be subdivided into two distinct subsets based on cell‐surface phenotype and tissue distribution.
Anti‐α4 integrin antibody induces apoptosis in murine thymocytes and staphylococcal enterotoxin B‐activated lymph node T cells
Authors: TCHILIAN* E. Z. , OWEN J. J. T. , JENKINSON E. J.
Published: November 2003

We have shown that an antibody (9C10) to the α4 integrin induces apoptosis in murine immature CD4+ CD8+ thymocytes and in activated (but not resting) mature lymph node T cells. In both cases, apoptosis is blocked by the highly selective protein kinase C (PKC) inhibitor Ro31.8425, suggesting that 9C10 induces signalling through the α4 integrin resulting in PKC activation leading to apoptosis. Overall, our results indicate the potential role of the α4 integrin‐mediated interactions in apoptosis induction during T‐cell development and following mature T‐cell activation.
Autocrine expression of interleukin‐7 rescues lymphoid expansion in interleukin‐7‐deficient mice
Authors: RICH B. E.
Published: November 2003

The murine interleukin‐7 (IL‐7) gene was disrupted to examine the role of IL‐7 in the lymphoid system. Expansion of lymphoid cells is sharply curtailed in IL‐7‐deficient mice. This is evident in a dramatic reduction but not elimination of lymphoid cells in the thymus, bone marrow and spleen. The few thymocytes present express CD4 and/or CD8 markers associated with T‐cell maturation. Similarly, a limited number of B cells detected in the bone marrow rearrange and express immunoglobulin genes. Small but distinct populations of B and T cells are found in the spleens of IL‐7‐deficient mice. Thus the signal transmitted by IL‐7 plays a central role in the expansion of lymphocytes while it is not absolutely required for their maturation. A transgene that directs expression of IL‐7 to lymphoid cells was found to restore the numbers of thymocytes, bone marrow B‐cell progenitors and splenic lymphocytes of IL‐7‐deficient mice to approximately normal levels. This genetic complementation confirms that the lymphoid defect is specifically due to the absence of IL‐7 and demonstrates that the expansion of lymphoid cells in an organism is regulated by their exposure to IL‐7.
Langerhans cells require signals from both tumour necrosis factor‐α and interleukin‐1β for migration
Authors: CUMBERBATCH M. , DEARMAN R. J. , KIMBER I.
Published: November 2003

The induction phase of contact sensitization is associated with the movement of epidermal Langerhans cells (LC) from the skin and their migration, via afferent lymphatics, to draining lymph nodes where they accumulate as immunostimulatory dendritic cells (DC). It has been demonstrated previously that tumour necrosis factor‐α (TNF‐α) provides an important signal for LC migration and that in the absence of this cytokine, movement of LC from the epidermis to regional lymph nodes is inhibited. Recent evidence indicates that interleukin‐1β (IL‐1β), a cytokine produced in murine epidermis exclusively by LC, may also play a role in LC migration. The purpose of the investigations described here was to clarify, using relevant neutralizing anti‐cytokine antibodies, the contributions made by TNF‐α and IL‐1β to the migration of LC from the epidermis. It was found that like anti‐TNF‐α, anti‐IL‐1β administered systemically to mice (by intraperitoneal injection), prior to skin sensitization with the contact allergen oxazolone, resulted in a marked inhibition of DC accumulation in draining lymph nodes. It was shown also that anti‐IL‐1β inhibited TNF‐α‐induced LC migration and DC accumulation and that, in similar fashion, the stimulation of LC migration and DC accumulation induced by IL‐1β was compromised by prior treatment with anti‐TNF‐α. Based upon these data it is proposed that the stimulation of LC migration in response to skin sensitization requires the receipt by LC of two independent signals, one provided by TNF‐α and the other by IL‐1β. Morphological analyses of LC in epidermal sheets prepared from animals exposed to these cytokines with or without prior systemic treatment with anti‐cytokine antibody suggested that the changes induced in LC by TNF‐α and IL‐1β may include the altered expression of adhesion molecules and acquisition of the ability to interact with and pass through the basement membrane.