bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased resistance to mycobacterial infection in the absence of interleukin‐10
Authors: Jacobs M. , Brown N. , Allie N. , Gulert R. , Ryffel B.
Published: March 2003

Interleukin‐10 (IL‐10) down‐regulates T helper type 1 cell and macrophage functions. As IL‐10 is induced along with tumour necrosis factor (TNF) and IL‐12 in mycobacterial infection, we asked whether endogenous IL‐10 plays a role in the antimycobacterial response. We demonstrate here that IL‐10‐deficient mice eliminate Mycobacterium bovis Calmette–Guérin bacillus faster than wild‐type mice. Granulomas are significantly larger, containing more CD‐11b‐ and CD11c‐positive antigen‐presenting cells and T cells, and the expression of major histocompatibility complex class II and intracellular adhesion molecule‐1 is increased. Macrophages in granulomas of IL‐10‐deficient mice express high levels of TNF, acid phosphatase and inducible nitric oxide synthase (iNOS). Finally, an increased cutaneous delayed‐type hypersensitivity reaction to mycobacterial proteins is further evidence of an augmented cell‐mediated immune response. In conclusion, the cell‐mediated immunity is enhanced in the absence of IL‐10, resulting in a robust granuloma response, which accelerates the clearance of mycobacteria. Therefore, endogenous IL‐10 attenuates mycobacterial immunity.
Intestinal lymphocyte number, migration and antibody secretion in young and old rats
Authors: Thoreux K. , Owen R. L. , Schmucker D. L.
Published: March 2003

This study demonstrates that the mucosal immune response to cholera toxin (CT) is compromised in old rats in comparison with young animals. The total number of immunoglobulin A (IgA)‐secreting cells is similar or higher in the intestinal inductor and effector sites in old animals. However, the number of specifically induced anti‐CT IgA antibody‐secreting cells is lower in these tissues in comparison with those in young animals. The kinetics of this immune response in the different gut‐associated lymphoid tissues studied suggests that the age‐associated decline in the number of anti‐CT IgA‐secreting cells in the intestinal mucosa reflects impaired IgA immunoblast migration. Our data from lymphocyte adoptive transfer studies indicate that factors intrinsic to both the donor cells and the host recipient influence the migration of immunoblasts from the Peyer's patches to the effector site. For example, donor cells from old donors transferred to either young or old recipient rats migrate slower than young donor lymphocytes transferred into old host animals. In vitro studies clearly indicate that ageing does not impair antibody secretion by intestinal mucosal plasma cells. Therefore, the age‐related decline in the intestinal mucosal immune response, e.g. diminished specific antibody titres in intestinal lavage, reflects fewer antibody‐secreting cells in the mucosa.
Major histocompatibility complex class II– fetal skin dendritic cells are potent accessory cells of polyclonal T‐cell responses
Authors: Elbe‐Bürger, A. , Mommaas A. M. , Prieschl, E. E. , Fiebiger, E. , Baumruker T. , Stingl G.
Published: March 2003

Whereas dendritic cells (DC) and Langerhans cells (LC) isolated from organs of adult individuals express surface major histocompatibility complex (MHC) class II antigens, DC lines generated from fetal murine skin, while capable of activating naive, allogeneic CD8+ T cells in a MHC class I‐restricted fashion, do not exhibit anti‐MHC class II surface reactivity and fail to stimulate the proliferation of naive, allogeneic CD4+ T cells. To test whether the CD45+ MHC class I+ CD80+ DC line 80/1 expresses incompetent, or fails to transcribe, MHC class II molecules, we performed biochemical and molecular studies using Western blot and polymerase chain reaction analysis. We found that 80/1 DC express MHC class II molecules neither at the protein nor at the transcriptional level. Ultrastructural examination of these cells revealed the presence of a LC‐like morphology with indented nuclei, active cytoplasm, intermediate filaments and dendritic processes. In contrast to adult LC, no LC‐specific cytoplasmic organelles (Birbeck granules) were present. Functionally, 80/1 DC in the presence, but not in the absence, of concanavalin A and anti‐T‐cell receptor monoclonal antibodies stimulated a vigorous proliferative response of naive CD4+ and CD8+ T cells. Furthermore, we found that the anti‐CD3‐induced stimulation of naive CD4+ and CD8+ T cells was critically dependent on the expression of FcγR on 80/1 DC and that the requirement for co‐stimulation depends on the intensity of T‐cell receptor signalling.
HLA‐G has a concentration‐dependent effect on the generation of an allo‐CTL response
Authors: Kapasi K. , Albert S. E. , Yie S.‐M. , Zavazava N. , Librach C. L.
Published: March 2003

Human leucocyte antigen (HLA) ‐G is expressed on trophoblast cells during pregnancy, suggesting a role in protection of the semiallogeneic fetus. Published data suggest that HLA‐G protects a cell against natural killer cell lysis. It has been hypothesized that HLA‐G may also protect the fetus by preventing allo‐cytotoxic T lymphocyte (CTL) responses. To test this hypothesis, we assayed the effects of various concentrations of purified HLA‐G on CTL response in a mixed lymphocyte culture (MLC) system. We found that concentrations ≥ 0·1 µg/ml of HLA‐G suppressed the allo‐CTL response by 30–100% over the control, but, paradoxically, concentrations of 0·01–0·05 µg/ml of HLA‐G augmented the allo‐CTL response by 25–50% over the control. Concentrations ≤ 0·001 µg/ml HLA‐G had no effect. Addition of HLA‐G to preprimed allo‐CTL effector cells did not affect their killing ability. Allo‐CTL suppressive doses of HLA‐G induced a T helper type 2 (Th2) cytokine response, whereas allo‐CTL‐enhancing doses of HLA‐G induced a Th1‐type cytokine response. HLA‐G purified from first‐trimester placenta does not affect allo‐proliferative responses nor does it alter the percentage of CD4+ or CD8+ T cells in MLCs. These findings support a potential role for HLA‐G‐mediated suppression of allo‐CTL formation in normal pregnancies. In addition, the effects observed at lower concentrations of HLA‐G may have interesting implications for the condition of pre‐eclampsia in which concentrations of this HLA class I molecule are reduced.
The calcium‐independent protein kinase C participates in an early process of CD3/CD28‐mediated induction of thymocyte apoptosis
Authors: Asada A. , Zhao Y. , Komano H. , Kuwata T. , Mukai M. , Fujita K. , Tozawa Y. , Iseki R. , Tian H. , Sato K. , Motegi Y. , Suzuki R. , Yokoyama M. , Iwata M.
Published: March 2003

Thymocyte negative selection eliminates self‐reactive clones and involves both a T‐cell receptor (TCR)/CD3‐mediated signal and a costimulatory signal, which can be delivered via CD28. Anti‐CD3/anti‐CD28‐triggered apoptosis in isolated CD4+CD8+ thymocytes in vitro provides a basic model for negative selection. Effects of isoform‐selective and non‐isoform‐selective inhibitors of protein kinase C (PKC) on this apoptotic process suggest that activation of Ca2+‐independent PKC isoforms during the first 2–3 hr of culture is essential for inducing apoptosis, and that Ca2+‐dependent PKC isoforms may be influential, but not essential, for apoptosis. To assess the CD3/CD28‐mediated activation of PKC in the apoptotic process, we prepared CD4+CD8+ thymocytes (without contamination with cells that had received negative or positive selection signals in vivo) by establishing TCR‐transgenic mice with RAG‐2‐deficient and non‐selecting major histocompatibility complex (MHC) backgrounds, in addition to a CD4+CD8+ thymocyte‐enriched population from normal mice. Translocation of Ca2+‐independent PKC from the cytosolic fraction to the particulate fraction of CD4+CD8+ thymocytes was induced by CD3/CD28‐mediated stimulation, but not by CD3‐ or CD28‐mediated stimulation alone, and peaked 2 hr after the start of culture. The kinase activity of the translocated Ca2+‐independent PKC was dependent on cofactors in vitro, indicating that novel (n)PKC, but not atypical (a)PKC or a proteolytic PKC fragment, was responsible for the activity. Immunoblotting analysis indicated that the nPKC‐θ isoform was the major contributor among nPKC isoforms, and that the classical (c)PKC‐α isoform was the major contributor among cPKC isoforms. These results suggest that activation of nPKC (especially the θ isoform) in CD4+CD8+ thymocytes is involved in a pathway for negative selection.
Induction of T helper 1‐ and T helper 2‐type immune responses duringHaemonchus contortus infection in sheep
Authors: Gill H. S. , Altmann K. , Cross M. L. , Husband A. J.
Published: March 2003

The production of cytokines by lymphoid cells, isolated from non‐infected and Haemonchus contortus‐infected lambs, was investigated. Particular attention was paid to differences in T helper 1‐ (Th1) and Th2‐type immune profiles between genetically resistant and random‐bred animal groups. Non‐infected resistant and random‐bred lambs produced equivalent levels of interferon‐γ (IFN‐γ) and interleukin‐5 (IL‐5), from isolated abomasal lymph node cells (ALN), mesenteric lymph node cells (MLN) and spleen cells (SC), in response to in vitro stimulation with T‐cell mitogen (concanavalin A) or larval parasite antigen. ALN and MLN cells derived from infected resistant and random‐bred lambs produced relatively lower levels of IFN‐γ, following in vitro stimulation with parasite antigen, when compared with their uninfected counterparts. In contrast, infected lambs of both groups showed enhanced mitogen‐ and antigen‐stimulated production of IL‐5, in comparison with uninfected controls, at days 5 and 28 postinfection (p.i.). Mitogen‐ and antigen‐stimulated IL‐5 responses were higher among resistant lambs compared with random‐bred lambs, with the highest overall production of IL‐5 by parasite antigen‐stimulated ALN and MLN cells. Among day 28 p.i. lambs, levels of cell culture‐derived parasite‐specific immunoglobulin G1 (IgG1) and IgE antibodies were higher in resistant lambs than in random‐bred lambs, following in vitro stimulation of SC or ALN cells with parasite antigen. Finally, after 28 days p.i., histological examination of abomasal tissue revealed higher densities of mast cells and eosinophils in the mucosa of resistant lambs than in random‐bred lambs. Taken together, these data support the notion of a strong Th2‐type immune response to Haemonchus infection in genetically resistant sheep, and support the claim for a Th1/Th2 dichotomy in ruminants.
Antigen receptor signalling in apoptosis‐resistant mutants of WEHI 231 cells
Authors: Tasker L. , Marshall‐clarke S.
Published: March 2003

Ligation of membrane immunoglobulin M (mIgM) induces cell cycle arrest and apoptosis in the WEHI 231 B‐lymphoma cell line. The molecular mechanisms which link receptor ligation and the nuclear events that underlie this response, have yet to be fully elucidated. Here we have examined the signals induced following mIgM cross‐linking in variants of WEHI 231 that no longer undergo apoptosis in response to this stimulus. Tyrosine phosphorylation of cellular substrates in two of the variants is identical to that seen in wild‐type cells but in one of the mutants, VS2.12, a restricted set of substrates becomes tyrosine phosphorylated. In a second variant (E8), mIgM cross‐linking does not induce elevation of intracellular Ca2+, although tyrosine phosphorylation of PLCγ2 is induced to an equivalent extent to that seen in WEHI 231 cells. A third variant, 2E10.F9, is resistant to apoptosis despite the fact that all signals analysed appear to be similar to those induced in wild‐type cells. Our findings show that resistance to apoptosis can arise as a result of mutations affecting discrete stages of the mIgM signalling pathway. The mutant lines reported here show defects that have not yet been identified in previous studies and are likely to be useful tools in dissecting the signalling of cell death in B lymphocytes.
Individual and combined effect of granulocyte–macrophage colony‐stimulating factor and prolactin on maturation of dendritic cells from blood monocytes under serum‐free conditions
Authors: Matera L. , Galetto A. , Geuna M. , Vekemans K. , Ricotti E. , Contarini M. , Moro F. , Basso G.
Published: March 2003

Prolactin (PRL) shares structural and functional features with haemopoietic factors and cytokine peptides. Dendritic cells (DC) are involved in both initiating the primary and boosting the secondary host immune response and can be differentiated in vitro from precursors under the effect of granulocyte–macrophage colony‐stimulating factor (GM‐CSF) plus other factors. Because PRL has been shown to functionally interact with GM‐CSF, we have addressed its role on GM‐CSF‐driven differentiation of DC. Monocytic DC precursors from peripheral blood mononuclear cells (PBMC) were enriched either by adhesion to a plastic surface or CD14‐positive selection and cultured for 7 days in serum‐free medium containing GM‐CSF, interleukin (IL)‐4 and PRL, alone or in combination. Cells with large, veiled cytoplasm, expressing major histocompatibility complex (MHC) class II and the costimulatory molecules CD80, CD86 and CD40 and lacking the monocyte marker CD14, were considered as having the phenotype of cytokine‐generated DC. Functional maturation was assessed by proliferation and interferon‐γ (IFN‐γ) release of allogeneic T lymphocytes. Physiological (10–20 ng/ml) concentrations of PRL interacted synergistically with GM‐CSF and the effect was similar to that induced by IL‐4 on GM‐CSF‐driven DC maturation. When used alone, the physiological concentrations of PRL were inhibitory, whereas higher concentrations (80 ng/ml) were stimulatory. The synergistic effect of PRL may in part be caused by its ability to counteract the down‐modulation of the GM‐CSF receptor observed in serum‐free conditions. These data provide further evidence of the significance of PRL in the process of T lymphocyte activation.
Differential cellular immunolocalization of renal tumour necrosis factor‐α production during ischaemia versus endotoxaemia
Authors: Donnahoo K. K. , Meng X. , Ao L. , Ayala A. , Shames B. D. , Cain M. P. , Harken A. H. , Meldrum D. R.
Published: March 2003

Both renal ischaemia and endotoxaemia provoke renal dysfunction and cellular injury. Although the clinical manifestation of each insult is similar (global renal dysfunction), ischaemia and endotoxaemia induce different patterns of cellular injury. Tumour necrosis factor‐α (TNF‐α) has been implicated in both types of renal injury; however, it remains unknown whether differential cellular TNF‐α expression accounts for these changes. We hypothesized that renal glomerular cells and tubular cells differentially express TNF‐α in response to ischaemia compared with endotoxaemia. To investigate this hypothesis, male Sprague–Dawley rats were anaesthetized and exposed to various time‐periods of renal ischaemia, with or without reperfusion (sham operation=negative control), or lipopolysaccharide (LPS) 0·5 mg/kg intraperitoneally (i.p.). The kidneys were harvested following renal injury, and rat TNF‐α protein expression was determined (by enzyme‐linked immunosorbent assay), as were TNF‐α bioactivity (by WEHI‐164 cell clone cytotoxicity assay) and TNF‐α cellular localization (by immunohistochemistry). TNF‐α protein expression and TNF‐α bioactivity peaked following 1 hr of ischaemia and 2 hr of reperfusion (48 ± 11 pg/mg of protein, P < 0·05, and 12 ± 0·5 × 10−3 units/mg of protein, P < 0·05, respectively). The concentration of TNF‐α increased to a similar extent following exposure to LPS; however, while TNF‐α production following ischaemia‐reperfusion injury localized predominantly to renal tubular epithelial cells, animals exposed to LPS demonstrated a primarily glomerular distribution of TNF‐α production. Hence, the cellular localization of renal TNF‐α production appears to be injury specific, i.e. renal tubular cells are the primary source of TNF‐α following an ischaemic insult, whereas LPS induces glomerular TNF‐α production. The cellular source of TNF‐α following different insults may have therapeutic implications for targeted inhibition of TNF‐α production.
Low concentrations of lipopolysaccharide synergize with peptides to augment human T‐cell proliferation and can prevent the induction of non‐responsiveness by CTLA4‐Ig
Authors: Goodier M. R. , Londei M.
Published: March 2003

We investigate how lipopolysaccharide (LPS) could influence antigen‐specific T‐cell responses as well as tolerance induction. Using the recall antigen tetanus toxoid for primary in vitro T‐cell stimulation, we observed that LPS synergized with peptides to augment proliferation, particularly when used at low concentrations (as little as 100 pg/ml), and that interleukin‐12 (IL‐12) was partially required for this synergistic effect. Because of the clear enhancement of in vitro peptide‐specific responses we then tested whether LPS could influence antigen‐specific tolerance driven by coincubation of antigen (tetanus toxoid; TT or immunodominant peptides) with human CTLA‐4Ig fusion protein. As expected, CTLA‐4Ig treatment inhibited responses to peptides. LPS (100 pg/ml) induced a partial recovery of primary in vitro proliferation under these conditions and the presence of LPS during the primary stimulation prevented the induction of tolerance normally observed on re‐stimulation with the same antigen alone. Contrary to the synergistic effects on peptide proliferation this action was not caused by release of IL‐12. In addition, the neutralization of tumour necrosis factor‐α (TNF‐α) during the primary stimulation did not inhibit proliferation on re‐stimulation with peptide. LPS could therefore exert dramatic effects on antigen‐specific proliferation and CTLA‐4Ig‐induced non‐responsiveness in human T cells, although via distinct mechanisms. These results reinforce the evidence that LPS influences T‐cell function, most likely as a consequence of myeloid cell activation.
Effect of early fetal splenectomy on prenatal B‐cell development in sheep
Authors: Press C. McL. , McCullagh P. , Landsverk T.
Published: March 2003

The contribution of early splenic B‐cell populations to the colonization of the ileal Peyer's patch was investigated following the surgical removal of the spleen in a series of 56‐day‐old fetal sheep. The fetuses were killed at 140 days of gestation and the ileal Peyer's patch, the distal jejunal lymph node which drains the Peyer's patch, and a peripheral lymph node, the superficial cervical lymph node, were examined. Enzyme and immunohistochemical evaluation concluded that the distribution of B cells, T cells and stromal cells in the ileal Peyer's patch was similar in splenectomized and normal fetal sheep. Thus, the presence of the fetal spleen was not essential for the colonization of the ileal Peyer's patch and other early sites of B‐cell accumulation would appear capable of generating the necessary precursor populations. Investigation of B‐cell populations in lymph nodes used a combination of terminal deoxynucleotidyl‐transferase‐mediated deoxyuridine‐triphosphate nick‐end‐labelling (TUNEL) histochemistry and immunofluorescence to determine the average number of apoptotic B cells in the primary follicles of the outer cortex of splenectomized and normal lambs. A significantly increased number of apoptotic B cells was present in the distal jejunal lymph node but not in the superficial cervical lymph node of splenectomized lambs. This finding suggests that splenectomy affected prenatal B‐cell development in fetal sheep and raises questions as to the regulation of B‐cell lymphopoiesis in a species using a post‐rearrangement organ of diversification.
The failure of Daudi cells to express the cellular prion protein is caused by a lack of glycosyl‐phosphatidylinositol anchor formation
Authors: Morelon E. , Dodelet V. , Lavery P. , Cashman N. R. , Loertscher R
Published: March 2003

The cellular prion protein (PrPc) is a glycosyl‐phosphatidylinositol (GPI)‐linked cell surface protein, which is expressed at high density on nervous tissues and at lower levels on most other solid‐organ tissues. It is also expressed on peripheral blood mononuclear cells (PBMC) of all lineages. In lymphocytes, its level of expression is dependent upon the state of cell activation, and polyclonal anti‐PrP antisera partially block lectin‐induced T‐cell activation, suggesting a functional role of the protein in this process. Using the monoclonal antibody (mAb) 3F4 we examined PrPc surface immunoreactivity on leukaemic cell lines of T‐ and B‐cell origin, and unexpectedly observed a complete lack of PrPc cell‐surface expression in Daudi cells, while all other cell lines displayed discernible reactivity. We demonstrated the intracellular presence of PrP‐specific mRNA and PrP protein. The lack of surface PrPc is unrelated to the well‐known defect of β2‐microglobulin (β2m) expression in Daudi cells as other β2m‐deficient cells, such as the melanoma cell line F0‐1 and spleen cells from β2m gene‐deleted mice, were not deficient in cell‐surface PrPc. Daudi cells failed to bind antibodies directed against all GPI‐linked cell surface proteins. In somatic hybridization experiments using murine spleen cells as partners, we observed de novo expression of human PrPc, CD55 and CD59, thus demonstrating in Daudi cells the availability of these gene products for GPI linkage and cell‐surface expression.
Immunological memory in B‐cell‐deficient mice conveys long‐lasting protection against genital tract infection with Chlamydia trachomatis by rapid recruitment of T cells
Authors: Johansson M. , Lycke N.
Published: March 2003

The role of antibodies and antigen deposition for the development of immunological memory has been incompletely investigated. We addressed whether long‐term protection and T‐cell memory can be stimulated against a genital tract infection with human Chlamydia trachomatis serovar D in B‐cell‐deficient (µMT) mice. At 6 months following a primary infection with C. trachomatis, both µMT and wild‐type (WT) mice exhibited strong and comparable protection against reinfection. Evidence of long‐lasting CD4+ T‐cell memory was found in both µMT and WT mice, typified by comparable delayed‐type hypersensitivity (DTH) reactions against chlamydial antigens. No bacterial or chlamydial DNA was found in the genital tract of µMT memory mice, suggesting that immunological memory was maintained in the absence of antigen. Whereas few T cells were present in the genital tract of memory mice, rapid recruitment of CD4+, and some CD8+, T cells into the genital tract tissue was observed after challenge with live bacteria. Accumulation of T cells in the genital tract was preceded by a short transient infection of similar magnitude in both µMT and WT memory mice, arguing against a long‐term protective role of local antibodies. The rapid recruitment of CD4+ T cells into the genital tract was associated with a transient detection of interferon‐γ (IFN‐γ) mRNA in the genital tract in chlamydia‐immune memory mice, which was not found in naïve, challenged mice. Thus, long‐term protection in the genital tract against C. trachomatis infection is conveyed by IFN‐γ‐producing CD4+ memory T cells, which appear to be maintained in the absence of antibodies and local antigen deposition.
Interleukin‐10 induces macrophage apoptosis and expression of CD16 (FcγRIII) whose engagement blocks the cell death programme and facilitates differentiation
Authors: Wang Z.‐Q. , Bapat A. S. , Rayanade R. J. , Dagtas A. S. , Hoffmann M. K.
Published: March 2003

The development of monocytes into macrophages is regulated by helper T cells (Th) cells and, vice versa, the differentiation of Th cells into Th1 and Th2 is regulated by macrophages. Herein we examined the role of the Th2 cytokine, interleukin‐10 (IL‐10), on the development of macrophages. IL‐10 is known to block the expression of antigen‐presenting major histocompatibility complex (MHC) II and of costimulatory B7 molecules but it induces the expression of FcRs, especially the FcγRIII (CD16). The expression of CD16 enables the macrophage to carry out antibody‐dependent cell‐mediated cytotoxicity (ADCC) functions. However, this differentiation step is largely undercut by the capacity of IL‐10 to induce macrophage apoptosis before the process of differentiation ensues. We found that the negative effect of IL‐10 on the survival of macrophages is reversed in an environment that contains immunoglobulin G (IgG). IgG, especially when immune complexed with antigen, stimulates CD16 to transmit survival signals in macrophages which enable them to complete the differentiation process into CD16+ cells. Thus, IL‐10 suppresses macrophage accumulation in healthy tissues where IgG is absent, and facilitates macrophage accumulation and differentiation in tissues that contain IgG, for example inflamed tissues or tissues that present autoreactive antibodies.
Nitric oxide and bone
Authors: Van'T Hof Rob J. , Ralston Stuart H.
Published: March 2003

Nitric oxide (NO) is a free radical which has important effects on bone cell function. The endothelial isoform of nitric oxide synthase (eNOS) is widely expressed in bone on a constitutive basis, whereas inducible NOS is only expressed in response to inflammatory stimuli. It is currently unclear whether neuronal NOS is expressed by bone cells. Pro‐inflammatory cytokines such as IL‐1 and TNF cause activation of the iNOS pathway in bone cells and NO derived from this pathway potentiates cytokine and inflammation induced bone loss. These actions of NO are relevant to the pathogenesis of osteoporosis in inflammatory diseases such as rheumatoid arthritis, which are characterized by increased NO production and cytokine activation. Interferon gamma is a particularly potent stimulator of NO production when combined with other cytokines, causing very high concentrations of NO to be produced. These high levels of NO inhibit bone resorption and formation and may act to suppress bone turnover in severe inflammation. The eNOS isoform seems to play a key role in regulating osteoblast activity and bone formation since eNOS knockout mice have osteoporosis due to defective bone formation. Other studies have indicated that the NO derived from the eNOS pathway acts as a mediator of the effects of oestrogen in bone. eNOS also mediates the effects of mechanical loading on the skeleton where it acts along with prostaglandins, to promote bone formation and suppress bone resorption. Pharmacological NO donors have been shown to increase bone mass in experimental animals and preliminary evidence suggests that these agents may also influence bone turnover in man. These data indicate that the l‐arginine/NO pathway represents a novel target for therapeutic intervention in the prevention and treatment of bone diseases.
Disrupted B‐lymphocyte development and survival in interleukin‐2‐deficient mice
Authors: Schultz Michael , Clarke Stephen H. , Arnold Larry W. , Sartor R. Balfour , Tonkonogy Susan L.
Published: March 2003

Interleukin‐2‐deficient (IL‐2−/−) mice develop a spontaneous, progressive, CD4+ T‐cell‐mediated colitis with an age‐related decrease in the number of B lymphocytes. The aim of this study was to determine the mechanisms of B‐cell loss in IL‐2−/− mice. Serum immunoglobulin G1 (IgG1) levels in 8‐week‐old IL‐2−/− mice were above normal but then decreased dramatically with advancing age. Between 8 and 11 weeks of age, the number of B‐cell progenitors (B220+ IgM−) in the bone marrow of IL‐2−/− mice was less than half of those in IL‐2+/+ littermates. By 22 weeks of age, very few progenitor cells remained in the bone marrow of most mice, and spleens were almost devoid of B cells. Likewise, B1 cells were not present in the peritoneal cavity of aged IL‐2−/− mice. Flow cytometry analysis of B‐cell differentiation in the bone marrow suggested a progressive loss of B cells from the most mature to the least mature stages, which was not dependent on IL‐2 receptor‐α (IL‐2Rα) expression. B cells transferred from normal animals had similar survival rates in IL‐2−/− and wild‐type mice. We conclude that conventional B cells in older IL‐2−/− mice are lost by attrition owing to a derangement in B‐cell development. Because B1 cells are less dependent on the bone marrow, a separate mechanism for their loss is suggested.
Differential modulation of CD8β by rat γδ and αβ T cells after activation
Authors: Straube Frank , Herrmann Thomas
Published: March 2003

Major histocompatibility complex (MHC) class I‐restricted αβ T cells express the CD8αβ heterodimer, which acts as a MHC class I‐specific co‐receptor. Rats are so far the only species with frequent expression of the CD8αβ by MHC‐unrestricted γδ T cells. This study compares CD8αβ expression by splenic rat αβ and γδ T cells and reveals a lineage‐specific difference in the control of CD8β expression. After activation in vitro, many γδ T cells, but not αβ T cells, persistently down‐modulate the expression of CD8β, but not CD8α, at the RNA level. Down‐regulation occurred after stimulation with T‐cell receptor (TCR)‐specific monoclonal antibody (mAb) and interleukin‐2 (IL‐2) or CD28‐mediated costimulation, and after activation with phorbol 12‐myristate 13‐acetate (PMA) and ionomycin. Functional differences between modulating and non‐modulating cells were not found with respect to interferon‐γ (IFN‐γ) production and cytolytic activity. The modulation could be indicative for a fundamental difference between αβ and γδ T cells and also limits the use of CD8β as a stable marker of γδ T‐cell subsets. Possibly, CD8β modulation provides a mechanism to escape over‐stimulation by (auto‐)antigens by increasing the threshold of TCR‐mediated activation in γδ T cells.
Interleukin‐1β, but not interleukin‐1α, is required for T‐cell‐dependent antibody production
Authors: Nakae Susumu , Asano Masahide , Horai Reiko , Iwakura Yoichiro
Published: March 2003

Interleukin‐1 (IL‐1) consists of two molecules, IL‐1α and IL‐1β, and IL‐1 receptor antagonist (IL‐1Ra) is a natural inhibitor of these molecules. Although the adjuvant effects of exogenously administered IL‐1 in the humoral immune response are well known, the roles of endogenous IL‐1 and the functional discrimination between IL‐1α and IL‐1β have not been elucidated completely. In this report, we investigated the role of IL‐1 in the humoral immune response using gene‐targeted mice. Both primary and secondary antibody production against T‐dependent antigen, sheep red blood cells (SRBC), was significantly reduced in IL‐1α/β−/− mice, and was enhanced in IL‐1Ra−/− mice. The intrinsic functions of B cells, such as antibody production against type 1 T‐independent antigen, trinitrophenyl–lipopolysaccharide and proliferative responses against mitogenic stimuli, were normal in IL‐1α/β−/− mice. The proliferative response of T cells and cytokine production upon stimulation with anti‐CD3 monoclonal antibody were also normal, as was the phagocytotic ability of antigen‐presenting cells (APCs). However, SRBC‐specific proliferative response and cytokine production of T cells through the interaction with APCs were markedly impaired in IL‐1α/β−/− mice, and enhanced in IL‐1Ra−/− mice. Moreover, we show that SRBC‐specific antibody production was reduced in IL‐1β−/− mice, but not in IL‐1α−/− mice. These results show that endogenous IL‐1β, but not IL‐1α, is involved in T‐cell‐dependent antibody production, and IL‐1 promotes the antigen‐specific T‐cell helper function through the T‐cell–APC interaction.
Interleukin‐10 promotes B16‐melanoma growth by inhibition of macrophage functions and induction of tumour and vascular cell proliferation
Authors: García‐Hernández M. L. , Hernández‐Pando R. , Gariglio P. , Berumen J.
Published: March 2003

The aim of this study was to investigate the mechanisms by which interleukin‐10 (IL‐10) induces tumour growth in a mouse‐melanoma model. A B16‐melanoma cell line (B16‐0) was transfected with IL‐10 cDNA and three clones that secreted high (B16‐10), medium and low amounts of IL‐10 were selected. Cell proliferation and IL‐10 production were compared in vitro, and tumour growth, percentages of necrotic areas, tumour cells positive for proliferating cell nuclear antigen (PCNA), IL‐10 receptor (IL‐10R) and major histocompatibility complex type I (MHC‐I) and II (MHC‐II), as well as infiltration of macrophages, CD4+ and CD8+ lymphocytes and blood vessels were compared in vivo among IL‐10‐transfected and non‐transfected tumours. Proliferation and tumour growth were greater for IL‐10‐transfected than for non‐transfected cells (P < 0·001), and correlated with IL‐10 concentration (r ≥ 0·79, P < 0·006). Percentages of tumour cells positive for PCNA and IL‐10R were 4·4‐ and 16·7‐fold higher, respectively, in B16‐10 than in B16‐0 tumours (P < 0·001). Macrophage distribution changed from a diffuse pattern in non‐transfected (6·4 ± 1·7%) to a peripheral pattern in IL‐10‐transfected (3·8 ± 1·7%) tumours. The percentage of CD4+ lymphocytes was 7·6 times higher in B16‐10 than in B16‐0 tumours (P = 0·002). The expression of MHC‐I molecules was present in all B16‐0 tumour cells and completely negative in B16–10 tumour cells. In B16‐0 tumours, 89 ± 4% of the whole tumour area was necrotic, whereas tumours produced by B16‐10 cells showed only 4·3 ± 6% of necrotic areas. IL‐10‐transfected tumours had 17‐fold more blood vessels than non‐transfected tumours (61·8 ± 8% versus 3·5 ± 1·7% blood vessels/tumour; P < 0·001). All the effects induced by IL‐10 were prevented in mice treated with a neutralizing anti‐IL‐10 monoclonal antibody. These data indicate that IL‐10 could induce tumour growth in this B16‐melanoma model by stimulation of tumour‐cell proliferation, angiogenesis and immunosuppression.
Liposome/DNA complexes coated with biodegradable PLA improve immune responses to plasmid encoding hepatitis B surface antigen
Authors: Bramwell Vincent W. , Eyles Jim E. , Somavarapu Satyanarayana , Alpar H. Oya
Published: March 2003

We hypothesized that the addition of polymer to the surface of liposome/DNA complexes may potentially enhance in vivo delivery of plasmid DNA to antigen‐presenting cells and thereby facilitate enhanced immune responses to encoded protein. BALB/c mice were immunized subcutaneously or intramuscularly three times with a total of 50 µg of the plasmid pRc/CMV‐HBs(S) (ayw subtype) encoding for the hepatitis B surface antigen. We measured transgene‐specific total immunoglobulin G (IgG), IgG2a, IgG2b and IgG1 antibody responses as well as splenocyte and T‐cell proliferation and cytokine production upon re‐stimulation following immunization. Modification of lipid/DNA complexes by the polymer precipitation method used here for the addition of poly(d,l‐lactic acid) was found to be consistently and significantly more effective than either unmodified liposomal DNA or naked DNA in eliciting transgene‐specific immune responses to plasmid‐encoded antigen when administered by the subcutaneous route. In addition, the polymer‐modified formulations delivered by this route were more effective than naked DNA delivered by the intramuscular route in inducing antibody responses (n=5, P<0·03). Our observations provide ‘proof of principle’ for the use of these multicomponent formulations, which offer potential for manipulation and increased transfection efficiency in vivo for the purposes of genetic immunization.
Evidence from twins for acquired cellular immune hyperactivity in type 1 diabetes
Authors: Petrovsky Nikolai , Kyvik Kirsten O. , Bonnevie‐Nielsen Vagn , Beck‐Nielsen Henning , Green Anders , Harrison Leonard C.
Published: March 2003

Type 1 diabetes has been associated with an increased frequency of activated T cells and T‐cell hyperactivity to non‐specific and disease‐specific stimuli including the islet autoantigen glutamic acid decarboxylase 65 (GAD). To address whether T‐cell hyperactivity is genetic or acquired we measured whole blood cytokines in vitro in response to GAD or tetanus in 18 identical twin pairs, nine discordant for type 1 diabetes. In addition, the activity of 2′, 5′ oligoadenylate synthetase (OAS) in blood mononuclear cells was measured as a marker of viral infection. Interleukin‐2 (IL‐2) basally and IL‐2 and interferon‐γ (IFN‐γ) in response to GAD, were detected more frequently and at higher levels in diabetic compared to non‐diabetic twins. IL‐10 was not different between groups. OAS activity was increased in diabetic compared to non‐diabetic twins and showed a correlation with basal IL‐2 and GAD‐stimulated IFN‐γ and IL‐10. These findings suggest that T‐cell hyperactivity in type 1 diabetes is an acquired trait and could reflect persisting virus expression.
Immunological basis of the development of necrotic lesions following Mycobacterium avium infection
Authors: Flórido Manuela , Cooper Andrea M. , Appelberg Rui
Published: March 2003

Normal C57BL/6 mice infected with 106 colony‐forming units of a highly virulent strain of Mycobacterium avium developed a progressive infection characterized by loss of T cells from the tissues and infiltration with high numbers of heavily infected macrophages. In contrast, when C57BL/6 mice were infected with 102 colony‐forming units of the same strain they retained T cells and T‐cell reactivity in the tissues, and granulomas evolved into large masses that, at 4 months of infection, exhibited central necrosis. The development of these necrotic lesions did not occur in nude mice, nor in mice genetically deficient in CD4, interleukin‐12 (IL‐12) p40, interferon‐γ (IFN‐γ) and CD40 and were reduced in mice deficient in CD54 or IL‐6. They were less numerous but bigger in mice deficient in IL‐10 or the inducible nitric oxide synthase, correlating with the increased resistance to mycobacterial proliferation of these strains as compared to control mice. The appearance of necrosis was not affected in mice deficient in CD8α, T‐cell receptor δ, tumour necrosis factor receptor p55, and perforin, nor was it affected in mice over‐expressing bcl2. The appearance of necrosis could be prevented by administering antibodies specific for CD4, IL‐12p40, or IFN‐γ from the second month of infection when organized granulomas were already found. Our results show that the immunological mediators involved in the induction of protective immunity are also major players in the immunopathology associated with mycobacteriosis.
Production of interleukin‐13 by human dendritic cells after stimulation with protein allergens is a key factor for induction of T helper 2 cytokines and is associated with activation of signal transducer and activator of transcription‐6
Authors: Bellinghausen Iris , Brand Pia , Böttcher Ingo , Klostermann Bettina , Knop Jürgen , Saloga Joachim
Published: March 2003

Dendritic cells (DC) are able to induce not only T helper 1 (Th1) but also Th2 immune responses after stimulation with allergens. While DC‐derived interleukin (IL)‐12 and IL‐18 are the key factors for the induction of Th1 cells, early signals being involved in Th2 differentiation are less well characterized so far. To analyse such early signals we used an antigen‐specific setting with CD4+ T cells from atopic donors stimulated in the presence of autologous mature DC, which were pulsed with different allergen doses. The addition of increasing amounts of allergen during DC maturation with tumour necrosis factor‐α, IL‐1β and prostaglandin E2 resulted in enhanced secretion of IL‐6 and IL‐12 by DC followed by increased production of Th1 (interferon‐γ; IFN‐γ) as well as Th2 (IL‐4, IL‐5) cytokines by CD4+ T cells. The coculture of allergen‐treated DC and CD4+ T cells also led to a dose‐dependent expression of active signal transducer and activator of transcription‐6 (STAT6), which was visible already after 1 hr. Additionally, rapid phosphorylation of STAT6 was seen in immature DC after stimulation with allergens but not with lipopolysaccharide or human serum albumin. STAT6 phosphorylation was associated with the production of IL‐13 by DC. The addition of neutralizing anti‐IL‐13 antibodies during maturation of DC inhibited STAT6 phosphorylation in CD4+ T cells as well as the production of IL‐4, and to a lesser extent of IL‐5, while IFN‐γ production was not affected. Addition of exogenous IL‐13 enhanced mainly the secretion of IL‐4. Taken together, DC‐derived IL‐13, which is released after exposure to allergens appears to be one of the critical factors for DC to acquire the capability to induce Th2 cytokine production.
Dendritic cells recruited to the lung shortly after intranasal delivery of Mycobacterium bovis BCG drive the primary immune response towards a type 1 cytokine production
Authors: Lagranderie Micheline , Nahori Marie‐Anne , Balazuc Anne‐Marie , Kiefer‐Biasizzo Hélène , Lapa E Silva Jose‐Roberto , Milon Geneviève , Marchal Gilles , Vargaftig Boris B.
Published: March 2003

We showed in a previous study that the intranasal (i.n) delivery of bacille Calmette–Guérin (BCG) to BP2 mice (H‐2q) inhibits eosinophilia and bronchial hyperreactivity in a mouse model of asthma. The present work has been performed to characterize the leucocyte lineages recruited to the lungs of mice after i.n. delivery of BCG and potentially involved in the polarization of T lymphocytes. The different antigen‐presenting cells (APC) recruited to bronchoalveolar lavage (BAL) and to lung tissue of mice shortly after the delivery of BCG were analysed in parallel as well as their capacity to drive the immune response towards a T helper type 1 cytokine production. Alveolar macrophages (AM) from the BAL were CD11c+, F4/80+ and CD11b−, and in the lung tissue two major populations of potential APC were detected: one CD11c−, F4/80+, CD11b+ and I‐Aq− was identified as interstitial macrophages (IM) and a second expressing CD11c+ and I‐Aq+ antigens, negative for CD11b and F4/80 markers as leucocytic dendritic cells (DC). Freshly isolated DC up‐regulated CD11b and CD40 antigens after overnight culture, but remained negative for CD8α antigen, suggesting a myeloid origin. Lung DC which produced high amount of interleukin (IL)‐12 were potent inducers of naive CD4+ T lymphocyte priming, as assessed by interferon‐γ (IFN‐γ) production by these naive CD4+ T cells. Lung explants recovered long term after BCG delivery produced sustained levels of IFN‐γ. Our results suggest that AM and particularly DC by secreting IL‐12 shortly after BCG delivery induce the long‐term persistence of IFN‐γ‐secreting T cells percolating in BCG‐loaded lung tissue.
Trust, power and interorganizational information systems: the case of the electronic trading community TransLease
Authors: Allen David K. , Colligan David , Finnie Andrew , Kern Thomas
Published: March 2003

Abstract.

This paper focuses on Cap Gemini’s electronic commerce system, TransLease. TransLease is an interorganizational information system (IOS), which facilitates electronic commerce between motor vehicle leasing and repair companies. During our investigation, the system was used by approximately 1000 repair agents working for seven of the UK’s leading vehicle leasing and contract hire companies. This system was originally developed by AT&T and acquired by Cap Gemini in July 1998. At the time of acquisition, the system was seen as being of high strategic value, although it was also seen as underperforming. This paper reports the results of an action research project, which formed one element of the process by which Cap Gemini investigated the former problem. In the paper, TransLease is described as a complex electronic community, dependent upon the existence of symbiotic relationships. As such, the problems that the system users and developers experienced can be attributed to factors that impeded the mutual benefit accruing from participation in the system. The efficacy of the terms of exchange and the degree to which participants mutually benefit through electronic interaction is determined by the complex interplay of a number of relational and organizational factors. The research therefore illustrates the importance of the ‘soft’ organizational issues in IOS management and development, and suggests a conceptual model of the factors relevant in this case. At the time of this study, TransLease was still in the early stages of its life cycle, having only been available in the marketplace for approximately 18 months. During this time, through recognizing the complex problems and issues detailed in this paper, Cap Gemini accordingly redressed the way in which the system was managed and maintained. TransLease is now seen as having matured into a highly successful example of an IOS – a view reflected by its position as market leader in the industry. As this paper will show, the key to improving the existing service has been the emphasis Cap Gemini now places on managing the ‘soft’ aspects of the electronic community.
‘It’s lots of bits of paper and ticks and post‐it notes and things . . .’: a case study of a rapid application development project
Authors: Beynon‐Davies Paul , Mackay Hugh , Tudhope Douglas
Published: March 2003

Abstract.

This paper reports an in‐depth case study of a rapid application development (RAD) project. RAD is a recent information systems development method noted for its high levels of user involvement and use of iterative prototyping. The paper develops a model of the core features of RAD gleaned from literature such as that published on the dynamic systems development method (DSDM). We report an ethnographic study of a RAD project and use this case material to contrast the theory with the practice of RAD. We conclude the paper with a consideration of a number of possible revisions to the prescriptions of RAD and also discuss the role of RAD within the broader context of IS development.

‘It’s lots of bits of paper and ticks and post‐it notes and things . . . you’ll get to understand it, it’s not that bad really’. A RAD team member
The significance of context in information systems and organizational change
Authors: Avgerou Chrisanthi
Published: March 2003

This paper argues that it is of crucial importance that information systems (IS) research and practice associates technology innovation with the context within which it is embedded. It identifies three principles to be followed in order to address the contextual processes involved in IS implementation: first, technology innovation should be considered in relation to socio‐organizational change; second, analysis should consider not only the local organizational, but also the national and international context; and third, analysis should consider both the technical/rational decisions and actions involved in the innovation process and the cultural, social and cognitive forces of such a process. These principles are demonstrated with the analysis of a case study of organizational reform in Cyprus.
Code quality analysis in open source software development
Authors: Stamelos Ioannis , Angelis Lefteris , Oikonomou Apostolos , Bleris Georgios L.
Published: March 2003

Abstract Proponents of open source style software development claim that better software is produced using this model compared with the traditional closed model. However, there is little empirical evidence in support of these claims. In this paper, we present the results of a pilot case study aiming: (a) to understand the implications of structural quality; and (b) to figure out the benefits of structural quality analysis of the code delivered by open source style development. To this end, we have measured quality characteristics of 100 applications written for Linux, using a software measurement tool, and compared the results with the industrial standard that is proposed by the tool. Another target of this case study was to investigate the issue of modularity in open source as this characteristic is being considered crucial by the proponents of open source for this type of software development. We have empirically assessed the relationship between the size of the application components and the delivered quality measured through user satisfaction. We have determined that, up to a certain extent, the average component size of an application is negatively related to the user satisfaction for this application.