bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of lisofylline and pentoxifylline on the bacterial‐stimulated production of TNF‐α, IL‐1β and IL‐10 by human leucocytes
Authors: VAN FURTH A. M. , VERHARD‐SEIJMONSBERGEN E. M. , VAN FURTH R. , LANGERMANS J. A. M.
Published: November 2003

The present study concerns the effect of the xanthine derivates lisofylline (LSF) and pentoxifylline (PTX) on the production of pro‐inflammatory cytokines tumour‐necrosis factor‐α (TNF‐α) and interleukin‐1β (IL‐1β) and the de‐activating cytokine interleukin‐10 (IL‐10) by human leucocytes during stimulation with lipopolysaccharide (LPS), heat‐killed Gram‐negative bacteria (GNB) or Gram‐positive bacteria (GPB). The production of TNF‐α and IL‐1β by leucocytes stimulated with LPS, Haemophilus influenzae type b (Hib) or Streptococcus pneumoniae was inhibited by both drugs. The production of IL‐10 by leucocytes stimulated with LPS and Hib was inhibited by both xanthine derivates only at 48 hr. However, incubation of leucocytes with S. pneumoniae in the presence of LSF or PTX stimulated the production of IL‐10 about four‐ and twofold at 24 hr and 48 hr, respectively. In all instances, the extent of inhibition or enhancement of cytokine production by LSF or PTX was equal. The divergent effects of xanthine derivates on the IL‐10 production indicate the existence of distinct intracellular pathways depending on whether leucocytes are stimulated by GPB or GNB.
Enhancement of antigen‐presenting ability of B lymphoma cells by partial inhibition of protein synthesis through inducing B7‐1 expression
Authors: AOI T. , NAKANO H. , TANAKA Y. , KAKIUCHI T.
Published: November 2003

During the investigation of the role of protein synthesis in antigen‐presenting cell (APC) function of A20‐HL B lymphoma cells, we found that partial inhibition of protein synthesis enhanced their APC function. The treatment of A20‐HL cells with 0·313–2·5 μm emetine, an irreversible inhibitor of protein synthesis, decreased protein synthesis by 60–70%, and enhanced their APC function to stimulate I‐Ad/OVA323–339‐specific T cells to produce interleukin‐2 in response to ovalbumin (OVA). The emetine‐treated and paraformaldehyde‐fixed A20‐HL cells required only 20 nm OVA323–339 peptide to stimulate the T cells, whereas those untreated and fixed required 200 nm peptide. This enhancement of APC function was mostly because of the induction of B7‐1 expression on A20‐HL cells by the emetine treatment, since B7‐1 molecules were detected on the emetine‐treated A20‐HL cells, but only negligibly, if at all, on the untreated cells, and an anti‐B7‐1 monoclonal antibody, 1G10, inhibited the enhanced APC function of the emetine–treated A20‐HL cells. The emetine‐treatment also increased B7–1 mRNA expression in A20‐HL cells, suggesting that the induction of B7‐1 expression was due to the increase in the accumulation of mRNA and the translation with residual ability to synthesize protein. Thus, partial inhibition of protein synthesis in A20‐HL cells increases B7‐1 mRNA accumulation and its expression on the cell surface, which results in the enhancement of their APC function.
The importance of MHC‐I and MHC‐II responses in vaccine efficacy against lethal herpes simplex virus type 1 challenge
Authors: GHIASI H. , ROOPENIAN D. C. , SLANINA S. , CAI S. , NESBURN A. B. , WECHSLER S. L.
Published: November 2003

To investigate the importance of major histocompatability complex (MHC) class I‐ and MHC class II‐dependent immune responses in herpes simplex virus‐1 (HSV‐1) vaccine efficacy, groups of β2m°/° (MHC I–) and Ab°/° (MHC II–) mice were inoculated with various vaccines, and then challenged intraperitoneally with HSV‐1. Following vaccination with either live avirulent HSV‐1, expressed HSV‐1 glycoprotein D (gD), or a mixture of seven expressed HSV‐1 glycoproteins (7gPs), Ab°/° (MHC‐II–) mice developed no enzyme‐linked immunosorbent assay (ELISA) or neutralizing antibody titres. In contrast, significant ELISA and neutralizing antibody titres were induced in β2m°/° (MHC‐I–) mice by all three vaccines. The neutralizing antibody titres were similar for all three vaccines, but were only ≈ 1/4 to 1/3 of that developed in C57BL/6 (parental) mice vaccinated with the same antigens. All three vaccines protected 100% of the wild‐type C57BL/6 mice against lethal challenge with 2×107 plaque‐forming units (PFU) of HSV‐1. The live virus vaccine and the 7gPs vaccine also protected 80% of the β2m°/° mice against the same lethal HSV‐1 challenge dose. In contrast, in Ab°/° mice, none of the vaccines provided significant protection against the same lethal challenge dose of HSV‐1. However, at a lower challenge dose of 2×106 PFU, all three vaccines protected 70–80% of the vaccinated Ab°/° mice (compared to only 10% survival in mock vaccinated controls). Thus, vaccination provided some protection against lethal HSV‐1 challenge in both β2m°/° and Ab°/° mice; however, the protection was less than that seen in the parental C57BL/6 mice. In addition, Ab°/° mice were less well protected by vaccination than were β2m°/° mice. Our results suggest that (1) both MHC‐I and MHC‐II are involved in vaccine efficacy against HSV‐1 challenge; (2) both types of responses must be present for maximum vaccine efficacy; and (3) the MHC‐II‐dependent immune response appeared to be more important than the MHC‐I‐dependent immune response for vaccine efficacy against HSV‐1 challenge.
Interferon‐regulatory factors during development of CD4 and CD8 thymocytes
Authors: SIMON A. K. , DESROIS M. , SCHMITT‐VERHULST A.‐M.
Published: November 2003

Selection events in the thymus occur at the double‐positive CD4+CD8+ (DP) developmental stage leading either to further differentiation of the CD4+ and CD8+ lineages or to deletion. The interferon‐regulatory factor IRF‐1 has been implicated in signalling for T‐cell death and also in CD8+ thymic differentiation. IRF‐1 is an activator and IRF‐2 a repressor of gene transcription regulated by type 1 interferons (IFN). Toevaluate the role of IRF‐1 and IRF‐2 in the differentiation of CD4 and CD8 thymocytes, we analysed their DNA‐binding activity before and after antigenic stimulation at different stages of thymic development and in peripheral T cells. Unseparated, double‐positive and single‐positive thymocytes as well as peripheral T lymphocytes from mice transgenic (tg) for a T‐cell receptor (TCR), restricted either by major histocompatibility complex class I or class II, were stimulated by their nominal antigen. Our results demonstrate that the DNA‐binding activity of IRF‐2 and, weakly, that of IRF‐1 are inducible in total thymocytes in response to antigen. There is no induction of IRF‐1/IRF‐2 binding activity at the double‐positive stage of thymic development in the MHC class II‐restricted model whereas in the MHC class I‐restricted model IRF‐1/IRF‐2 activity is induced weakly. At the single‐positive stage, antigen induces the IRF‐1/IRF‐2 DNA binding in both CD4+ and CD8+ thymocytes, but not in mature lymphocytes from the periphery. This pattern of expression suggests that IRF‐1/IRF‐2 binding activities resulting from antigen stimulation are developmentally regulated. No evidence for a selective role of IRF‐1 in the development of the CD8+ lineage was found, however.
An essential role for endogenous interferon‐γ in the generation of protective T cells against Mycobacterium bovis BCG in mice
Authors: YANG J. , MITSUYAMA M.
Published: November 2003

Protective CD4+ T cells against Mycobacterium bovis bacillus Calmette–Guérin(BCG), which are characterized by the ability to produce interferon‐γ (IFN‐γ), could be induced by immunization of mice with viable BCG but not with killed BCG. A high level of IFN‐γ mRNA was observed when normal spleen cells were stimulated with viable but not killed BCG. By comparing mice immunized with either viable or killed M. bovis BCG, it was found that a high level of IFN‐γ mRNA was expressed only after immunization with viable BCG. This finding prompted us to investigate the effect of neutralizing the IFN‐γ on the final generation of protective T cells against M. bovis BCG. When endogenous IFN‐γ was neutralized by administration of anti‐IFN‐γ monoclonal antibody in mice immunized with viable BCG, the generation of protective T cells was significantly impaired, as revealed by the adoptive transfer of spleen T cells. The generation of BCG‐specific, IFN‐γ‐producing T cells was also abolished. These results clearly demonstrate that endogenous IFN‐γ actually plays a critical role in the generation of protective T cells against M. bovis BCG in vivo. Moreover, this study suggests that the lack of IFN‐γ‐inducing ability is responsible for the inability of killed M. bovis BCG to induce protective T cells in mice.
Distribution of cyclophilin B‐binding sites in the subsets of human peripheral blood lymphocytes
Authors: DENYS A. , ALLAIN F. , FOXWELL B. , SPIK G.
Published: November 2003

Cyclophilin B (CyPB) is a cyclosporin A (CsA)‐binding protein, mainly associated with the secretory pathway and released in biological fluids. We have recently demonstrated that both free CyPB and CyPB–CsA complex specifically bind to peripheral blood T lymphocytes and are internalized. These results suggest that CyPB might promote the targeting of the drug into sensitive cells. Peripheral blood lymphocytes are subdivided in several populations according to their biological functions and sensitivity to CsA. We have investigated the binding of CyPB to these different subsets using a CyPB derivatized by fluorescein through its single cysteine which retains its binding properties. We have confirmed that only T cells were involved in the interaction with CyPB. The ligand binding was found to be heterogeneously distributed on the different T‐cell subsets and surface‐bound CyPB was mainly associated with the CD4‐positive cells. No significant difference was noted between the CD45RA and CD45RO subsets, demonstrating that CyPB‐binding sites were equally distributed between native and memory T cells. CD3 stimulation of T lymphocytes led to a decrease in the CyPB‐binding capacity, that may be explained by a down‐regulation of the CyPB‐receptor expression upon T‐cell activation. Finally, we demonstrated that CyPB‐receptor‐positive cells, isolated on CyPB sulphydryl‐coupled affinity matrices, are more sensitive to CyPB‐complexed CsA than mixed peripheral blood lymphocytes, suggesting that CyPB potentiates CsA activity through the binding of the complex. Taken together, our results demonstrate that CyPB‐binding sites are mainly associated with resting cells of the helper T lymphocyte, and that CyPB might modulate the distribution of CsA through the drug targeting to sensitive cells.
Immunization with gastric H+/K+‐ATPase induces a reversible autoimmune gastritis
Authors: SCARFF K. J. , PETTITT J. M. , VAN DRIEL I. R. , GLEESON P. A. , TOH B.‐H.
Published: November 2003

The gastric H+/K+‐ATPase has been implicated as a major autoantigen in pernicious anaemia in humans and in thymectomy‐induced autoimmune gastritis in mice. Here we have shown that autoimmune gastritis can be generated by direct immunization of non‐thymectomized BALB/c mice with mouse gastric H+/K+‐ATPase in complete Freund’s adjuvant. The gastritis was characterized by infiltration of the gastric submucosa and mucosa with macrophages, CD4+ and CD8+ T cells, and B cells and by circulating autoantibodies to the H+/K+‐ATPase. The mononuclear infiltrate within the gastric mucosa was accompanied by loss of parietal and zymogenic cells and accumulation of small immature epithelial cells. Splenocytes from gastritic mice adoptively transferred gastritis to naive recipients. Cessation of immunization resulted in decrease in autoantibody titre and regeneration of parietal and zymogenic cells. The results directly confirm that the gastric H+/K+‐ATPase is the causative autoantigen in the genesis of autoimmune gastritis. Recovery of the lesion following cessation of immunization suggests that homeostatic mechanisms can reverse a destructive autoimmune process.
Macrophage migration inhibitory factor induces phagocytosis of foreign particles by macrophages in autocrine and paracrine fashion
Authors: ONODERA S. , SUZUKI K. , MATSUNO T. , KANEDA K. , TAKAGI M. , NISHIHIRA J.
Published: November 2003

Exposure to foreign particles sometimes causes inflammatory reactions through production of cytokines and chemoattractants by phagocytic cells. In this study, we focused on macrophage migration inhibitory factor (MIF) to evaluate its pathophysiological role in the phagocytic process. Immunohistochemical analysis of human pseudosynovial tissues retrieved at revision of total hip arthroplasty showed that infiltrating mononuclear and multinuclear cells were positively stained by both an anti‐CD68 antibody and anti‐human MIF antibody. For in vitro study, MIF was released from murine macrophage‐like cells (RAW 264.7) in response to phagocytosis of fluorescent‐latex beads in a particle dose‐dependent manner. Northern blot analysis showed marked elevation of the MIF mRNA level in the phagocytic macrophage‐like cells. Moreover, pretreatment of RAW 264.7 cells with rat recombinant MIF increased the extent of phagocytosis by 1·6‐fold compared with the control. Taken together, these results suggest that MIF plays an important role by activating macrophages in autocrine and paracrine fashion to phagocytose foreign particles.
Interferon‐γ receptor‐deficient mice exhibit impaired gut mucosal immune responses but intact oral tolerance
Authors: KJERRULF M. , GRDIC D. , EKMAN L. , SCHÖN K. , VAJDY M. , LYCKE N. Y.
Published: November 2003

Interferon‐γ (IFN‐γ) receptor knock‐out (IFN‐γR−/−) mice were used to analyse the role of IFN‐γ in mucosal immune responses following oral immunization. We found that the IFN‐γR−/− mice demonstrated 50% reduced spot‐forming cell (SFC) responses in the gut lamina propria and spleen after oral immunization with keyhole limpet haemocyanin (KLH) plus cholera toxin (CT) adjuvant. The IFN‐γR−/− mice exhibited 10‐fold reduced total serum KLH‐specific antibody levels compared with wild‐type mice after oral immunization, while after intravenous immunization, no such difference was seen, suggesting a selective impairment of mucosal immune responses. Moreover, oral immunizations resulted in impaired interleukin‐4 (IL‐4), IL‐10 and IFN‐γ production by spleen T cells from IFN‐γR−/− mice, indicating that no reciprocal up‐regulation of Th2‐activities had occurred despite the lack of IFN‐γR function. No reduction in Th1 or Th2 cytokines was observed following systemic immunizations. Despite potentially strong modulating effects of IFN‐γ on epithelial cell IgA transcytosis and electrolyte barrier functions, CT‐immunized IFN‐γR−/− mice demonstrated unaltered protection against CT in ligated intestinal loops together with normal anti‐CT IgA and total IgA levels in gut lavage. Oral feeding with KLH followed by parenteral immunization resulted in strongly suppressed SFC numbers and reduced cell‐mediated immunity in both wild‐type and IFNγR−/− mice. CT‐adjuvant abrogated induction of oral tolerance in both IFN‐γR−/− and wild‐type mice. Collectively, our data argue that the two major response patterns induced by oral administration of protein antigen, i.e. active IgA immunity and oral tolerance, are differently regulated. Thus, IFN‐γR−/− mice have impaired mucosal immune responses while induction of oral tolerance appears to be unaffected by the lack of IFN‐γ functions.
An antibody specific for interleukin‐6 reverses age‐associated changes in spontaneous and induced cytokine production in mice
Authors: GORCZYNSKI R. M. , CINADER B. , RAMAKRISHNA V. , TERZIOGLU E. , WAELLI TH. , WESTPHAL O.
Published: November 2003

A number of quantitative and qualitative changes in the pattern of cytokine production have been reported to accompany the process of ageing in laboratory animals and in human populations, including an increase in serum levels of interleukin‐1 (IL‐1) and IL‐6, as well as increased concanavalin A (ConA) – stimulated production of IL‐4, IL‐10 and transforming growth factor‐β (TGF‐β), and decreased production of IL‐2 from cultured spleen cells. Increased IL‐1 and IL‐6 production is a feature of splenic adherent cells and peritoneal exudate cells taken from aged mice and stimulated with lipopolysaccharide in vitro. We have asked whether the altered production of lymphocyte‐derived cytokines (IL‐4, IL‐2, TGF‐β) is itself a function of a primary alteration in IL‐1/IL‐6 production (from macrophage/monocytes) by infusing monoclonal antibodies to these cytokines prior to harvesting cells from aged mice and stimulating the cells in vitro. Anti‐IL‐6, but not anti‐IL‐1, reversed the age‐associated alteration in lymphocyte cytokine production. The general pattern of cytokine production in aged mice is of a type‐2 cytokine type, and thus these data are consistent with the idea that increased production of IL‐6 in aged animals is causally implicated in this age‐associated polarization to type‐2 cytokine production.
L‐selectin activates JNK via src‐like tyrosine kinases and the small G‐protein Rac
Authors: BRENNER B. , WEINMANN S. , GRASSMÉ H. , LANG F. , LINDERKAMP O. , GULBINS E.
Published: November 2003

Selectin and &agr;4&bgr;7‐integrins have been shown to mediate transient leucocyte interactions with endothelial cells which is a crucial step in the initial immune response to pathogens. We have previously shown that stimulation of T lymphocytes via L‐selectin results in activation of a signalling cascade from the L‐selectin molecule via the tyrosine kinase p56lck and tyrosine phosphorylation of L‐selectin to the stimulation of p21Ras and Rac proteins. In the present study we demonstrate that stimulation of Jurkat T lymphocytes via L‐selectin results in an activation of Jun N‐terminal kinase (JNK) but not of p38‐K. L‐selectin‐initiated activation of JNK is mediated by src‐like tyrosine kinases and the small G‐protein Rac 1/2, since genetic or pharmacological inhibition of p56lck or Rac proteins prevent the stimulation of JNK by L‐selectin. Thus, the data point to a novel signalling cascade from L‐selectin via src‐like tyrosine kinases and Rac proteins to JNK.
n‐Butyrate downregulates the stimulatory function of peripheral blood‐derived antigen‐presenting cells: a potential mechanism for modulating T‐cell responses by short‐chain fatty acids
Authors: BÖHMIG G. A. , KRIEGER P‐M. , SÄEMANN M. D. , WENHARDT C. , POHANKA E. , ZLABINGER G. J.
Published: November 2003

Modulation of proliferative T‐cell responses by n‐butyrate has been suggested to result from direct interference with cell cycle progression. Considering the important role of antigen‐presenting cells (APC) in T‐cell activation, we were particularly interested in studying the impact of n‐butyrate on these cells. We demonstrated that pretreatment of human peripheral blood mononuclear cells (PBMC) or monocytes with this agent resulted in a dose‐ and time‐dependent downregulation of their capability to stimulate T‐cell responses with a similar pattern of inhibition when this agent was present throughout the culture period. Pretreatment with n‐butyrate was effective in preventing both alloresponses and T‐cell proliferation to immobilized anti‐CD3 monoclonal antibody (mAb) suggesting alteration of costimulatory function. Flow cytometric analysis revealed that interferon‐&ggr; (IFN‐&ggr;)‐induced upregulation of B7‐1 expression on monocytes was profoundly inhibited by n‐butyrate. Furthermore, this agent significantly suppressed the expression of intercellular adhesion molecule‐1 (ICAM‐1) or lymphocyte function‐associated antigen‐3 (LFA‐3). In contrast, constitutive as well as cytokine‐induced expression of B7‐2 was enhanced by n‐butyrate. Additionally, in monocytes, but not in T cells, treatment with n‐butyrate led to significant alteration of membrane integrity owing to apoptotic cell death. Our findings indicate that modulation of T‐cell responses by n‐butyrate could also result from altered APC function, possibly as a consequence of downregulating distinct adhesion and/or costimulatory receptors as well as of inducing apoptosis. A potential clinical relevance of short‐chain fatty acids for reducing T‐cell‐mediated immune reactions via modulating APC function is speculated.
Induction of selective anergy by toxic shock syndrome toxin‐1 in CD8+ T cells
Authors: ZHAO† Y.‐X. , HOLMDAHL R. , TARKOWSKI A.
Published: November 2003

Interferon‐&ggr; (IFN‐&ggr;) mRNA expression of murine splenic CD8+ T cells in response to toxic shock syndrome toxin‐1 (TSST‐1) was examined in vitro. In primary response to TSST‐1, purified CD8+ cells expressed a low frequency of IFN‐&ggr; mRNA and produced very little interleukin‐2 (IL‐2) compared with unseparated lymphocytes and purified CD4+ cells. The addition of IL‐2 to the cell culture enhanced IFN‐&ggr; mRNA expression in the CD8+ population. Upon restimulation with TSST‐1, no IL‐2 production and hardly any IFN‐&ggr; mRNA expression were observed in the purified CD8+ lymphoblasts. Exogenously added IL‐2 did not increase the IFN‐&ggr; mRNA expression in purified CD8+ blasts exposed to TSST‐1. In contrast, stimulation with concanavalin A induced a high frequency of IFN‐&ggr; mRNA in purified CD8+ blasts. Moreover, TSST‐1 induced strong IFN‐&ggr; mRNA expression in unseparated T lymphoblasts and purified CD4+ populations. Interestingly, a combined immunocytochemistry and in situ hybridization technique showed a high frequency of CD8+ IFN‐&ggr; mRNA‐expressing cells in an unseparated blast population. These results demonstrate that TSST‐1 induces a selective anergy within the purified CD8+ T‐cell compartment and that the IFN‐&ggr; mRNA expression in CD8+ T blasts can be achieved by stimulation of unseparated T cells with TSST‐1.
Presentation of cryptococcal capsular polysaccharide (GXM) on activated antigen‐presenting cells inhibits the T‐suppressor response and enhances delayed‐type hypersensitivity and survival
Authors: BLACKSTOCK R. , CASADEVALL A.
Published: November 2003

A hallmark of infection with Cryptococcus neoformans is depression of the immune system characterized by poor inflammatory responses and loss of delayed‐type hypersensitivity (DTH) and antibody responses. T‐suppressor cell (Ts) responses, elicited by the capsular polysaccharide (GXM) of the organism, are known to develop during infection. This study was undertaken to develop a method to inhibit the anti‐GXM Ts response and thereby study the influence of the Ts response on immune responsiveness and survival in cryptococcosis. Antigen‐presenting cells (APC), elicited with complete Freund’s adjuvant (CFA), were treated in vitro with GXM (GXM–APC). The GXM–APC were injected intravenously into normal mice. These mice were resistant to induction of anti‐GXM Ts cells when soluble GXM was administered in tolerogenic doses or when animals were infected with C. neoformans. Inhibition of the anti‐GXM Ts response was specific to GXM as levan–APC did not inhibit induction of anti‐GXM Ts cells. Inhibition of the anti‐GXM Ts response could not be attributed to increased clearance of GXM due to induction of anti‐GXM antibodies or other mechanisms. Anti‐cryptococcal DTH responses were lost in mice by the second week of infection. However, treatment with GXM–APC, but not levan–APC, allowed mice to maintain their DTH response. GXM–APC pretreatment enhanced survival of infected mice compared with mice pretreated with levan–APC. These results show that GXM–APC induces immune responses that inhibit the induction of Ts responses and enhances DTH responses in infected mice. These responses correlate with enhanced survival after cryptococcal infection.
Increased natural killer resistance to cyclosporine A by continuous doses of dexamethasone in rats
Authors: KAMIO E. , SAKAMOTO Y. , KIMURA T. , TAKAKUWA H. , SAKURAI‡ M. , HOSOKAWA J.‐I. , NAKAGAWA K. , YOSHIDA F. , TAGAMI K.
Published: November 2003

There is a controversy on the effects of physiological levels of glucocorticoids on natural killer (NK) cytotoxity. Therefore, the effects of exogenously administered dexamethasone on NK cytotoxity in 8‐week‐old male, Fischer 344 rats were studied. We suppose that the reason for the controversy is insufficient sensitivity of the ordinal radioactive chromium‐release assay for normal healthy subjects or animals. Therefore, we developed a new index, a resistance to artificial immunosuppressor, cyclosporine A (CsA) using rat NK activity as an indicator, and named this index, increased resistance to immunosuppressor (IRIS). After some basic, characterizing studies, authors confirmed the fact that continuous doses of dexamethasone (DEX) attenuated NK suppression of CsA. In protocol 4, 18 rats were randomly divided into three groups: the first (DEX+CsA) was injected for 5 days with 0·1 mg DEX/kg/day and a single dose of CsA on the final day, intraperitoneally; the second (SAL+CsA) was treated with an equal volume of saline and CsA; the third (DEX+SAL) was treated with DEX but not CsA. The IRIS in NK activity was increased significantly (P<0·01) with 5 days injection of DEX. These results demonstrated that physiological, and continuous dosage of glucocorticoids stimulated IRIS in NK activity in rats, and this suggests that appropriate stimuli through the hypothalamic–adrenal axis might be acting, at least, as a defence against immune collapses or dysfunctions.
A comparative analysis of cytokine production and tolerance induction by bacterial lipopeptides, lipopolysaccharides and Staphyloccocus aureus in human monocytes
Authors: KREUTZ M. , ACKERMANN U. , HAUSCHILDT S. , KRAUSE S. W. , RIEDEL D. , BESSLER W. , ANDREESEN R.
Published: November 2003

Monocytes (MO) and macrophages (MAC) are important producers of cytokines involved in the pathophysiology of bacterial sepsis. Most studies concentrate on the effects of bacterial lipopolysaccharides (LPS) regarding the induction of cytokine gene expression and secretion in MO/MAC. Here we report that besides LPS, the synthetic lipoprotein analogue lipopeptide N‐palmitoyl‐S‐(2,3‐bis(palmitoyl)‐(2RS)‐propyl)‐(R)‐cysteinyl‐alanyl‐glycine (Pam3‐Cys‐Ala‐Gly), another component of the outer membrane of Gram‐negative bacteria, as well as heat‐killed Staphyloccocus aureus (S. aureus/SAC) are potent stimuli for cytokines in human MO. For all three investigated stimuli we found an individual pattern of cytokine induction: LPS was most potent in inducing interleukin‐6 (IL‐6) synthesis, whereas for tumour necrosis factor‐α (TNF‐α) secretion SAC was the best stimulus. Comparable amounts of IL‐8 were induced by either LPS or Pam3‐Cys‐Ala‐Gly, with SAC being less effective even at higher concentrations. The addition of serum led to an increase in LPS‐, SAC‐ and Pam3‐Cys‐Ala‐Gly‐stimulated TNF‐α secretion, indicating that the presence of serum is critical not just for LPS stimulation. Furthermore, as is known for LPS, Pam3‐Cys‐Ala‐Gly and SAC rendered MO refractory to a second bacterial stimulus. Pam3‐Cys‐Ala‐Gly and SAC induced tolerance for itself, but LPS could partially overcome this effect. As the CD14 molecule is discussed as a common receptor for different bacterial components, we investigated whether the TNF‐α response of MO could be blocked by anti‐CD14 antibodies. MY4, a CD14 antibody, selectively blocked the TNF‐α secretion induced by LPS but not by Pam3‐Cys‐Ala‐Gly or SAC. In summary, we conclude that besides LPS, lipopeptide Pam3‐Cys‐Ala‐Gly and SAC are potent stimuli for human MO, while the mechanisms of activation seem to be partially different from LPS.
The defective antigen‐presenting activity of murine fetal macrophage cell lines
Authors: KHALILI H. , DESHPANDE R. , CHANG M‐d. Y.
Published: November 2003

We have previously reported that placental macrophages of fetal origin have a decreased ability to present antigen. To clarify the underlying mechanism for this deficiency, we have generated primary fetal macrophage cell lines. Our data show that despite their defective antigen‐presenting ability, fetal macrophages do express all known accessory molecules, intracellular adhesion molecule‐1, B7 and major histocompatibility complex class II molecules. However, fetal macrophages do not express detectable invariant chain mRNA which is known to have a major role in the class II‐associated antigen‐processing pathway. Since fetal macrophages can neither present antigenic peptides nor superantigen, the diminished invariant chain expression alone cannot account for the impaired antigen‐presenting function of fetal macrophages.
CD4 regulates the efficiency of an endogenous superantigen‐induced clonal deletion of TCRVβ11+ cells in the periphery
Authors: KUBO S. , HASEGAWA A. , HASHIMOTO K. , SHIMIZU C. , KUBO M. , TADA T. , NAKAYAMA T.
Published: November 2003

Peripheral T‐cell antigen receptor Vβ (TCRVβ) repertoire is influenced by clonal deletion both in the thymus and periphery. Developing thymocytes expressing certain TCRVβ are deleted by endogenous superantigens presented on major histocompatibility complex (MHC) molecules in the thymus. Likewise, mature T cells bearing particular TCRVβ chains can be clonally deleted by superantigens in the periphery. The efficiency with which T cells expressing particular Vβ subunits are deleted differs depending upon which coreceptor is expressed. Indeed, while deletion of Vβ11+ splenic T cells in CBA/J (Mls‐1,a I‐E,+ MTV 9+) mice is quite efficient for CD4+ spleen T cells, it is much less efficient for CD8+ splenic T cells. If the difference in the efficiency of deletion is due solely to the coreceptor expressed, then a transgene encoding CD4 should increase the efficiency with which CD8+ cells are deleted. To address this question, we have produced CD4 transgenic (TG) mice that express physiologic levels of CD4 on all thymocytes and peripheral CD8 T cells. CD4 molecules expressed on CD8+ splenic T cells were associated with p56lck tyrosine kinase, and were functional as evidenced by their ability to facilitate class II alloreactivity. Furthermore, we found that ectopic expression of TG CD4 molecules on CD8+ T cells was able to affect the efficiency of deletion in response to superantigen stimulation. In particular, deletion of TCRVβ11+ T cells was much less efficient for CD8+ than for CD4+ T‐cell subpopulations in (CBA/J×B6) F1 mice. However, expression of the CD4 transgene on CD8+ splenic T cells from these mice increased the efficiency of deletion in the CD8+ Vβ11 T cells. Interestingly, this effect was not observed in a mature CD8+ thymocyte subpopulation. The results in this report demonstrate that CD4 molecules are involved in peripheral deletion of TCRVβ11+ T cells in (CBA/J×B6) F1 mice, and that the TCRVβ repertoire can be altered by ectopic expression of CD4 on all T‐lineage cells.
Endothelial production of MCP‐1: modulation by heparin and consequences for mononuclear cell activation
Authors: DOUGLAS M. S. , ALI S. , RIX D. A. , ZHANG J.‐G. , KIRBY J. A.
Published: November 2003

Heparin is a polyanionic glycosaminoglycan (GAG) that can bind with high affinity to a range of cytokines including interferon‐γ (IFN‐γ) and members of the chemokine superfamily. This GAG also possesses immunomodulatory activity in vivo and can antagonize the capacity of IFN‐γ to induce class II MHC antigen expression, and to up‐regulate intercellular adhesion molecule‐1, by cultured endothelial cells. Previous studies have shown that binding to cell‐surface heparan sulphate is essential for optimal activity of IFN‐γ and that free heparin competitively inhibits this sequestration process. The present study was performed to increase our understanding of the immunosuppressive activity of heparin by investigation of potential antagonism of the production and function of monocyte chemotactic peptide‐1 (MCP‐1), a chemokine important for mononuclear leucocyte recruitment across vascular endothelium. It was found that mixture of heparin with IFN‐γ inhibited up‐regulation of the signal transducer and activator of transcription protein, STAT‐1 produced normally by treatment of endothelial cells with IFN‐γ. An inhibition of MCP‐1 production was observed that was specifically caused by mixture of IFN‐γ with heparin‐like, and therefore cytokine‐binding, GAGs. It was also shown that mixture of heparin‐like GAGs with MCP‐1 inhibited the rapid tyrosine phosphorylation of phosphatidylinositol 3‐kinase which is normally produced by treatment of mononuclear leucocytes with this chemokine. Blockade of this intracellular signalling event was associated with a reduction in the normal transendothelial migration response towards MCP‐1. Results from this study indicate that soluble, heparin‐like GAGs can block IFN‐γ‐dependent up‐regulation of MCP‐1 production by cultured endothelial cells, and can also antagonize the leucocyte‐activating and migration‐promoting properties of pre‐existing MCP‐1. These activities may contribute to the immunomodulatory properties of heparin.
Interleukin‐7 treatment promotes the differentiation pathway of T‐cell‐receptor‐αβ cells selectively to the CD8+ cell lineage
Authors: VARAS A. , VICENTE A. , JIMÉNEZ E. , ALONSO L. , MORENO J. , MUÑOZ J. J. , ZAPATA A. G.
Published: November 2003

In this report we have studied the influence of interleukin‐7 (IL‐7) on thymocyte differentiation by evaluating the effects of IL‐7 on the generation of T‐cell receptor‐αβ (TCR‐αβ) and TCR‐γδ thymocyte subpopulations in rat fetal thymus organ culture. IL‐7 enhanced the differentiation pathway of TCRαβ thymocytes, first increasing the numbers of immature CD8+ cells, and later those of both CD4+CD8+ and mature thymocytes. The kinetics of thymocyte migration out of thymic lobes was also accelerated, and the average number of mature TCR‐αβhi emigrants per day was increased in the presence of IL‐7. Moreover, mature CD4−CD8+ thymocytes were preferentially generated after IL‐7 administration. This TCR‐αβhi cell population was not actively dividing, indicating that IL‐7‐promoted thymocyte differentiation was selective to the CD8 cell lineage. Distribution of some TCR‐Vα and TCR‐Vβ segments among mature thymocytes was also modified in IL‐7‐treated thymic lobes. On the contrary, the maturation of TCR‐γδ was not affected by IL‐7 addition during the first days of culture, but their numbers sharply increased by day 6 of culture. These results were confirmed with IL‐7‐treated cultures for 24 hr, showing that IL‐7 responsiveness was acquired by TCR‐γδ cells late in thymus ontogeny. The present results thus indicate a key role for IL‐7 in the maturation of TCR‐αβ thymocytes and the expansion of thymic TCR‐γδ cells.
Locomotor properties of human germinal centre B cells: activation by anti‐CD40 and IL‐4 allows chemoattraction by anti‐immunoglobulin
Authors: KOMAI‐KOMA M. , WILKINSON P. C.
Published: November 2003

The locomotor properties of B cells isolated from the germinal centres (GC) of human tonsils were studied using polarization, collagen gel invasion and micropore filter assays. The proportion of motile GC cells in the freshly isolated population was small. During culture in interleukin‐4 (IL‐4)+anti‐CD40, but not in control medium, the proportion of polarized cells increased and these cells migrated actively into collagen gels. After 24 hr culture, most of the surviving population was in locomotor morphology. The locomotor population consisted mainly of centrocytes in the G1 phase of growth. More locomotor cells than spherical cells took up []>
3H]uridine, but locomotor cells did not take up [3H]thymidine. After culture for 6 hr in IL‐4+anti‐CD40, GC B cells were tested in short‐term polarization assays and filter assays for their response to chemoattractants. In both assays, a proportion of the cells responded to anti‐IgA and to anti‐IgA F(ab′)2 fragments at 1 ng/ml, or to anti‐IgG, anti‐IgM and F(ab′)2 fragments of these antibodies at 100 ng–1 μg/ml. A checkerboard filter assay showed a good chemokinetic response and a weaker chemotactic response of GC cells to anti‐IgA. Expression of FcγRII (CD32) was increased after culture in IL‐4+anti‐CD40, and these cultured cells responded in filter and polarization assays to anti‐CD32. Thus culture in IL‐4 and anti‐CD40 not only rescues GC B cells, but also increases their locomotor capacity and allows them to respond in chemotaxis assays to anti‐immunoglobulin.
A comparative study of the in vitro immunomodulatory activity of human intact immunoglobulin (7S IVIG), F(ab′)2 fragments (5S IVIG) and Fc fragments. Evidence for post‐transcriptional IL‐2 modulation
Authors: NACHBAUR D. , HEROLD M. , EIBL B. , GLASSL H. , SCHWAIGHOFER H. , HUBER C. , GÄCHTER A. , PICHL M. , NIEDERWIESER D.
Published: November 2003

During the past few decades intravenous immunoglobulin (IVIG) has been used successfully in the treatment of various immunoregulatory disorders. Treatment results have been attributed to immunomodulation mainly via Fc receptors or by anti‐idiotypic antibodies to disease‐causing autoantibodies. From the present study it is clearly evident that 7S IVIG (intact immunoglobulin) as well as 5S IVIG [ F(ab′)2 fragments] and Fc fragments have a potent immunomodulatory capacity. We demonstrate that mainly 7S IVIG inhibits alloantigen‐induced T‐cell proliferation and generation of cytotoxic T lymphocytes. Reduced interleukin‐2 (IL‐2) protein levels in culture supernatants of IVIG‐supplemented mixed lymphocyte reactions (MLR) but unchanged IL‐2 mRNA levels strongly argue in favour of a post‐transcriptional interference of IVIG with cytokines and/or cytokine production. Interferon‐γ (IFN‐γ), soluble IL‐2 receptor (sIL‐2R) and monokines such as IL‐1β, IL‐6, IFN‐α and tumour necrosis factor (TNF‐α) were not affected by IVIG supplementation to MLR. Fc fragments were superior to F(ab′)2‐containing IVIG (5S and 7S IVIG) in inhibiting lectin stimulation of peripheral blood mononuclear cells (PBMC), whereas natural killer (NK) cytotoxicity was primarily inhibited by Fc‐bearing IVIG (7S IVIG and Fc fragments), suggesting multiple mechanisms of IVIG immunomodulatory activity.
A comparison of the antigen‐presenting capabilities of class II MHC‐expressing human lung epithelial and endothelial cells
Authors: CUNNINGHAM A. C. , ZHANG J.‐G. , MOY J. V. , ALI S. , KIRBY J. A.
Published: November 2003

Human lung alveolar epithelial cells constitutively express class II major histocompatibility complex (MHC). Human lung microvascular endothelial and small airway epithelial cells can be induced to express class II MHC by stimulation with the pro‐inflammatory cytokine interferon‐γ. The levels of class II MHC on lung epithelial and endothelial cells were comparable to those seen on an Epstein–Barr virus (EBV)‐transformed B‐cell line. However, the costimulatory molecules B7‐1 and B7‐2 were not expressed. The ability of the class II MHC expressing human lung parenchymal cells to present alloantigen to CD4+ T lymphocytes was investigated. Freshly isolated human alveolar epithelial cells (type II pneumocytes) and monolayers of interferon‐γ‐stimulated small airway epithelial and lung microvascular endothelial cells were co‐cultured with allogeneic CD4+ T lymphocytes and proliferation determined by []>
3H]thymidine incorporation. A clear difference was observed between effects of the epithelial and endothelial cells on CD4+ T‐lymphocyte activation. Alveolar and small airway epithelial cells failed to stimulate the proliferation of allogeneic CD4+ T lymphocytes whereas lung microvascular endothelial cells did stimulate proliferation. This difference could not be explained by the levels of class II MHC or the lack of B7‐1 and B7‐2 solely. Microvascular endothelial cells, and not alveolar or small airway epithelial cells, possess B7‐independent costimulatory pathways.
Inhibitors of dipeptidyl peptidase IV induce secretion of transforming growth factor‐β1 in PWM‐stimulated PBMC and T cells
Authors: REINHOLD D. , BANK U. , BÜHLING F. , LENDECKEL U. , FAUST J. , NEUBERT K. , ANSORGE S.
Published: November 2003

Various studies have shown that the membrane ectoenzyme dipeptidyl peptidase IV (DPIV; CD26), expressed on T, natural killer (NK) and B cells in the immune system, is involved in the regulation of DNA synthesis and cytokine production. We show that the specific DP IV inhibitors Lys[ Z(NO2)]‐thiazolidide, Lys[Z(NO2)]‐piperidide, and Lys[Z(NO2)]‐pyrrolidide inhibit DNA synthesis as well as production of interleukin‐2 (IL‐2), IL‐10, IL‐12, and interferon‐γ (IFN‐γ) of pokeweed mitogen (PWM)‐stimulated purified T cells. Most importantly, these inhibitors induce a three‐ to fourfold increased secretion of latent transforming growth factor‐β1 (TGF‐β1) by PWM‐stimulated peripheral blood mononuclear cells (PBMC) and T cells, as measured with a specific TGF‐β1 enzyme‐linked immunosorbent assay and in the Mv1Lu bioassay. As we could demonstrate previously, TGF‐β1 exhibits the same inhibitory effects as DP IV inhibitors on DNA synthesis and cytokine production (Cytokine 1994, 6, 382–8; J Interferon Cytokine Res 1995, 15, 685–90). A neutralizing chicken anti‐TGF‐β1 antibody was capable of abolishing the DP IV inhibitor‐induced suppression of DNA synthesis of PWM‐stimulated PBMC and T cells. These data suggest that TGF‐β1 might have key functions in the molecular action of DP IV/CD26 in regulation of DNA synthesis and cytokine production.
Role of E‐ and P‐selectin in migration of monocytes and polymorphonuclear leucocytes to cytokine and chemoattractant‐induced cutaneous inflammation in the rat
Authors: WALTER U. M. , ISSEKUTZ A. C.
Published: November 2003

The contribution of E‐ and P‐selectin in the rat to the migration of polymorphonuclear leucocytes (PMNL) and monocytes to acute dermal inflammation induced by a chemoattractant (C5ades Arg) or endothelial cell activating agents [ lipopolysaccharide, tumour necrosis factor‐&agr; (TNF‐&agr;), &agr;‐thrombin and interferon‐&ggr;] administered intradermally was investigated. Migration was quantitated using radiolabelled blood PMNL and monocytes and new, function‐blocking monoclonal antibodies (mAbs) to rat E‐ and P‐selectin were employed. Monocyte migration to inflamed skin was partially inhibited (40–75%) by P‐selectin mAb with all stimuli tested, but not by anti‐E‐selectin. PMNL migration in response to all stimuli was also inhibited (50–75%) by blocking P‐selectin, but in contrast to monocytes, PMNL accumulation was partially inhibited by mAb to E‐selectin in &agr;‐thrombin and TNF‐&agr; lesions. When P‐selectin was blocked by mAb, mAb to E‐selectin significantly inhibited further (by 70–90%) both PMNL and monocyte accumulation in all lesions, indicating that both P‐ and E‐selectin contribute to the migration of these leucocytes. Blocking L‐selectin in addition to P‐ and E‐selectin, had no effect on the remaining migration. Thus, optimal PMNL and monocyte migration to chemotactic factor‐ and cytokine‐induced skin inflammation is P‐selectin dependent. E‐selectin becomes important, in most conditions used here, when P‐selectin mediated recruitment is not operative. A selectin independent pathway likely mediates up to 20% of PMNL and monocyte migration to acute inflammation, at least in skin.
Formalized systems development methodologies: a critical perspective
Authors: Fitzgerald Brian
Published: November 2003

Systems development issues occupy a position of central importance in the information systems field and much has been prescribed in the quest for successful systems development. However, given the well‐documented ‘software crisis’, success is far from guaranteed for systems development projects. Many researchers see the solution to the software crisis in terms of increased control and the more widespread adoption of rigorous and formalized system development methodologies. This paper presents some arguments and pressures which support the use of methodologies and some evidence of the literature bias which favours methodologies is provided. However the problems associated with the use of methodologies have not perhaps received as much attention in previous research. This paper identifies a number of arguments and pressures which question the value of methodologies, and reports the results of a field study which investigated the role of methodologies in practice. The critical perspective adopted throughout shows that, contrary to the predominant view in literature, the assumption that increased adoption of methodologies addresses the problems inherent in systems development is by no means proven.
The fetish of technique: methodology as a social defence
Authors: Wastell David G.
Published: November 2003

Methodology is a central issue in the theory and practise of information systems development. Structured methods, for instance, have been widely championed as providing a way of improving the quality of software systems. In this paper a case study is presented in which a mail order company made an attempt to implement a well‐known methodology, Structured Systems Analysis and Design Methodology (SSADM). Far from facilitating the development process, SSADM encouraged a rigid and mechanical approach in which the methodology was applied in a ritualistic way which inhibited creative thinking. The argument is thus, that methodology, although its influence may be benign, has the potential to operate as a ‘social defence’, i.e. as a set of organizational rituals with the primary function of containing anxiety. The grandiose illusion of an all‐powerful method allows practitioners to deny their feelings of impotence in the face of the daunting technical and political challenges of systems development. By withdrawing into this fantasy world the learning processes that are critical to the success of systems development are jeopardized. Methodology, whilst masquerading as the epitome of rationality, may thus operate as an irrational ritual, the enactment of which provides designers with a feeling of security and efficiency at the expense of real engagement with the task at hand.
Stakeholders, soft systems and technology: separation and mediation in the analysis of information system requirements
Authors: Vidgen Richard
Published: November 2003

This paper describes the application of stakeholder analysis and soft systems thinking for an investigation of information system requirements. It is argued that it is appropriate to approach IS development as an exercise in managing complexity (soft systems) and pluralism (stakeholder analysis). A framework for investigating IS requirements is proposed that contrasts the current situation with the future situation and the real world with conceptual thinking about the real world. These aspects are viewed as outcomes of the process of IS requirements analysis, for which the metaphor of mediation is adopted in preference to presenting requirements analysis as a binary distinction between social construction and objectivity. The IS requirements analysis framework is applied in action research and the findings and learning that arose are presented.
Too close for comfort? Distance and engagement in interpretive information systems research
Authors: Nandhakumar Joe , Jones Matthew
Published: November 2003

In this paper, we seek to explore the implications of the assumptions underlying interpretivism for the preferred relationship between the researcher and the research phenomenon. The growing interest in interpretive information systems research has drawn attention to the need to gain access to the interpretations of social actors. Various data‐gathering methods are available to the researcher to achieve such access. These may be seen as located on a spectrum in terms of the degree of engagement between the researcher and the research subject. While engagement is not without its drawbacks from a research perspective, it may be argued that it enables good access to the sorts of data that interpretive researchers are seeking. Most of the research reported in the information systems (IS) field, however, has adopted relatively distant methods by which the researcher avoids intervention in the research context. Some of the characteristics of more engaged forms of research are illustrated through a discussion of a participant observation study of executive information systems development. This research has highlighted a number of issues that can be seen to be common to other data‐gathering methods. Reasons for the neglect of engaged data‐gathering methods are explored, and it is suggested that consideration of the issues it raises may contribute to more reflexive interpretive IS research practice.