bluuebunny/crossref_metadata_2025 huggingface dataset:
The role of the Fgr tyrosine kinase in the control of the adhesive properties of U937 monoblastoid cells and their derivatives
Authors: FAULKNER‡ L. , PATEL M. , BRICKELL§ P. M. , KATZ D. R.
Published: November 2003

In humans, expression of the cellular proto‐oncogene c‐fgr is normally restricted to mature cells of the myeloid lineage, mantle zone B cells and various myeloid and B‐cell lines. Previous studies of the monoblastoid cell line, U937, showed that c‐fgr expression increased following differentiation, but its role in monocytes and related cells has not been defined in functional terms. We therefore investigated the role of c‐fgr in U937 cells transfected with the c‐fgr gene such that its expression could be manipulated independent of differentiation. Induction of the transfected c‐fgr gene by cadmium ions did not affect cell proliferation, responses to phorbol 12‐myristate 13‐acetate (PMA), dihydroxycholecalciferol (DHCC), tumour necrosis factor‐α (TNF‐α) or retinoic acid, or phagocytosis of antibody‐coated sheep red blood cells. However, there was increased surface expression of CD54 (intracellular adhesion molecule‐1; ICAM‐1) and CD102 (ICAM‐2) and decreased surface expression of CD50 (ICAM‐3) compared with cells that had been transfected with plasmid only and treated in the same way. These findings suggest that the product of the c‐fgr gene may be important in control of relative adhesive properties of mature monocytic cells.
Differential display analysis of murine collagen‐induced arthritis: cloning of the cDNA‐encoding murine ATPase inhibitor
Authors: YAMADA E. , ISHIGURO N. , MIYAISHI O. , TAKEUCHI A. , NAKASHIMA I. , IWATA H. , ISOBE K.
Published: November 2003

We used the differential display technique in order to detect a new gene involved in murine type II collagen‐induced arthritis (CIA). In this study, we have identified a novel gene, IF1, whose expression level is increased during the natural course of CIA. Northern blot analyses suggest that IF1 is involved in the natural course of CIA but is not involved as a trigger of CIA. IF1 is considered to be the murine ATPase inhibitor gene for several reasons. First, IF1 shows an extremely high homology to the rat ATPase inhibitor; the highly conserved region between rat and bovine amino acid residues 22–45, which is the minimum sequence showing ATPase inhibitory activities, is also highly conserved in IF1. Second, IF1 possesses a histidine‐rich region in the same area, which is thought to be important for regulation of mammalian inhibitors. Third, the tissue distribution of IF1 is very suggestive. The expression of IF1 was very strong in energetic organs such as the heart, brain and kidney, and the development of arthritis requires great amounts of ATP. As arthritis develops rapidly, the cellular ATP pool may be decreased. Before the ATP pool is exhausted, the ATPase inhibitor may serve as a brake for ATP hydrolysis. If the supply of free energy can be reduced, the inflammation of arthritis may in turn be restored. Our hypothesis is that the ATPase inhibitor is involved in regulating the inflammatory responses.
Alternatively activated macrophages actively inhibit proliferation of peripheral blood lymphocytes and CD4+ T cells in vitro
Authors: SCHEBESCH C. , KODELJA V. , MÜLLER C. , HAKIJ N. , BISSON S. , ORFANOS C. E. , GOERDT S.
Published: November 2003

We compared the immunological functions of interferon‐γ (IFN‐γ)‐induced, classically activated macrophages (caMΦ) and of interleukin‐4 (IL‐4)‐ and glucocorticoid‐induced, alternatively activated macrophages (aaMΦ) in a human co‐culture system in vitro. Proliferation of peripheral blood leucocytes (PBL) or CD4+ T cells mediated by optimal doses of phytohaemagglutinin (PHA) or concanavalin A (Con A) was only marginally influenced by caMΦ, but was strongly inhibited by aaMΦ. The degree of lymphocyte proliferation sustained in the presence of caMΦ was gradually reduced in a dose‐dependent fashion by the addition of aaMΦ. Flow cytometric analysis revealed that expression of costimulatory molecules such as CD11a, CD40, CD54, CD58, CD80 and CD86 did not vary significantly between caMΦ and aaMΦ and was low for CD58, CD80 and CD86. As shown by reverse transcriptase‐polymerase chain reaction (RT‐PCR) analysis, IL‐10 was expressed in caMΦ, aaMΦ and control macrophages; the level of expression of IL‐10 was slightly enhanced in aaMΦ. Neither neutralizing anti‐IL‐10 antibodies, indomethacin nor NG‐monomethyl‐l‐arginine (NMMLA) was able to reverse aaMΦ‐mediated inhibition of lymphocyte proliferation. Of several agents interfering with various second messenger pathways, cAMP and the Ca2+‐ionophor A23187 inhibited differentiation of cultured human monocytes into phenotypically mature aaMΦ expressing MS‐1 high molecular weight protein (MS‐1‐HMWP) and RM 3/1 antigen, and prevented the suppressive action of aaMΦ on lymphocyte proliferation. In conclusion, these results show that aaMΦ actively inhibit mitogen‐mediated proliferation of PBL and CD4+ T cells independently of the expression of costimulatory molecules and of IL‐10, NO or prostaglandin synthesis, and that inhibition of phenotypic differentiation of aaMΦ is paralleled by a lack of functional maturation. Thus, fully matured aaMΦ may be functional in down‐regulating CD4+ T‐cell‐mediated immune reactions by an as yet unknown mechanism.
β2 (CD18) and β1 (CD29) integrin mechanisms in migration of human polymorphonuclear leucocytes and monocytes through lung fibroblast barriers: shared and distinct mechanisms
Authors: SHANG X. ‐Z. , ISSEKUTZ A. C.
Published: November 2003

Accumulation of leucocytes in inflamed lung tissue and alveolar space involves their migration through vascular endothelium and then lung connective tissue. As a model of this process, we investigated human polymorphonuclear leucocyte (PMNL) and monocyte migration through a biological barrier of human lung fibroblasts (HLF) grown on polycarbonate filters. Very few PMNL (1–2%) or monocytes (3–8%) migrated through the HLF barriers spontaneously. Migration increased to 48–53% of added PMNL and 17–24% of added monocytes, when a C5a chemotactic gradient was present. The monocyte migration induced by C5a was not inhibited by monoclonal antibodies (mAb) to CD18 (β2 integrins). This CD18‐independent migration was partially inhibited (35%) by mAb to α5 of VLA‐5 and completely inhibited by the combination of mAb to α4 of VLA‐4 with mAb to VLA‐5, in the presence of mAb to CD18. In contrast, PMNL migration across HLF induced by C5a was partially inhibited by mAb to CD18 alone, but even with the addition of mAb to VLA‐4,VLA‐5 and VLA‐6, the greatest degree of inhibition was only 60%. Blocking the function of CD18 was not required to observe the inhibition by mAb to VLA‐4, although the inhibitory effect of mAb to VLA‐5 and VLA‐6 alone or in combination was only observed when CD18 mechanisms were also blocked with anti‐CD18 mAb. These results demonstrate that (a) both monocytes and PMNL can use either CD11/CD18 (β2 integrin) or β1 (CD49/CD29) integrins to migrate through HLF barriers; (b) in the case of monocytes, the VLA‐4 and VLA‐5 β1 integrins account for essentially all the CD11/CD18‐independent migration mechanisms; and (c) in contrast to monocytes, PMNL CD18‐independent migration is mediated not only by VLA‐4 and VLA‐5, but also by VLA‐6, and up to 40% of the migration appears to be via yet to be defined PMNL surface molecules.
The repertoire of HLA–Cw‐specific NK cell receptors CD158 a/b (EB6 and GL183) in individuals with different HLA phenotypes
Authors: FROHN C. , SCHLENKE P. , KIRCHNER H.
Published: November 2003

Over the last few years, natural killer (NK) cells have been shown to express major histocompatibility complex (MHC) molecules recognizing receptors that are thought to function primarily as negative signalling receptors. Much attention has been focused on the NK cell receptors CD158a (EB6) and CD158b (Gl 183), which recognize two alternative epitopes on the HLA‐Cw locus. In order to investigate whether HLA type affects the CD158a/b repertoire, expressed as percentage positive cells for a particular receptor and mean expression on this population of NK cells, peripheral blood lymphocytes of 47 HLA‐typed donors were examined. Peripheral blood samples were examined by flow cytometric analysis to investigate the expression of CD158a and CD158b receptors on the surface of NK cells. In parallel, we determined each individual’s HLA phenotype. There was a great heterogeneity in CD158 expression; nevertheless all individuals had NK cells belonging either to the CD158 a+b−, a−b+ or a−b− populations. No positive or inverse correlations could be shown between either receptor expression intensity or proportion of positive cells, and presence of the appropriate ligand. Thus no association between an individual’s NK receptor repertoire and HLA serotype could be demonstrated. It is concluded that CD158 is expressed on NK cells in a highly redundant fashion. Our data do not support either a positive selection mechanism or the receptor calibration model.
Role of annexins in endocytosis of antigens in immature human dendritic cells
Authors: LARSSON M. , MAJEED M. , ERNST J. D. , MAGNUSSON K.‐E. , STENDAHL O. , FORSUM U.
Published: November 2003

We have evaluated the uptake of a soluble protein antigen, denitrophenylated human serum albumin (DNP‐HSA), and two different intracellular bacteria; Chlamydia trachomatis serovar L2 and Mycobacterium tuberculosis strain H37Ra, by immature human dendritic cells. These were generated by culturing progenitor cells from blood in the presence of cytokines (granulocyte–macrophage colony‐stimulating factor and interleukin‐4). Dendritic cells play a crucial part in antigen presentation for the induction of T‐cell‐dependent immune responses in various tissues. Recently, macropinocytic and phagocytic activity has been shown for immature dendritic cells of mouse, rat and human origin. In the present study, macropinocytosis characterized the uptake of the soluble protein‐antigen DNP‐HSA, whereas the C. trachomatis were ingested via receptor‐mediated endocytosis in coated pits, and opsonized M. tuberculosis via phagocytosis. To follow the intracellular routes of the antigens, their positions were compared with the localization of annexins, a family of Ca2+‐and phospholipid‐binding proteins, involved in membrane fusion, aggregation and transport of different vesicles. To elucidate further the intracellular pathway of the antigens, two other proteins, lysosome‐associated membrane protein‐1 (LAMP‐1) and cathepsin D, were labelled. They are known to colocalize with major histocompatibility complex class II compartments in the immature dendritic cells. We observed a distinct translocation of annexin V to DNP‐HSA containing endosomes, and annexin III to vesicles with C. trachomatis. Furthermore, annexin III, IV and V redistributed to phagosomes with M. tuberculosis. Both LAMP‐1 and cathepsin D colocalized with DNP‐HSA endosomes, and with phagosomes with M. tuberculosis. Thus, immature human dendritic cells have the capacity to phagocytose. Moreover, the handling of these antigens by dendritic cells may represent three distinct intracellular pathways, albeit some properties and compartments are shared.
B‐cell‐deficient mice develop complete immune protection against genital tract infection with Chlamydia trachomatis
Authors: JOHANSSON M. , WARD M. , LYCKE N.
Published: November 2003

We evaluated the ability of mice made genetically deficient for B cells to resolve a primary infection and to develop protective immunity against vaginal challenge with a human isolate of Chlamydia trachomatis bacteria. The B‐cell‐deficient μmT mice cleared a primary ascending infection with similar or faster kinetics compared with wild‐type mice. The presence of chlamydial inclusion bodies and the degree of inflammation in the upper genital tract was comparable and showed similar kinetics in μmT as in wild‐type mice. Following resolution of the primary infection the mice were challenged by 100 ID50 of live bacteria and the level of protection and the extent of local inflammation was assessed. Strikingly, all μmT mice, as well as most of the wild‐type mice, demonstrated complete immune protection with no bacterial shedding. While high titres of chlamydia‐specific antibodies were stimulated locally and systemically in wild‐type mice, no antibodies were detected in μmT mice. However, in both strains, immunohistochemical analysis of the upper genital tract demonstrated the presence of large numbers of CD4+ T cells and increased levels of interferon‐γ (IFN‐γ)‐producing cells. The results unequivocally demonstrate that antibodies are not required for full protection to develop against ascending infection with a high dose of C. trachomatis in the female genital tract. Our study confirms the notion that cell‐mediated immunity, in particular that owing to CD4+ T helper 1 (Th1)‐type cells, is critical for host resistance against C. trachomatis in mice.
Sequential development of intraepithelial γδ and αβ T lymphocytes expressing CD8αβ in neonatal rat intestine: requirement for the thymus
Authors: HELGELAND L. , BRANDTZAEG P. , ROLSTAD B. , VAAGE J. T.
Published: November 2003

Previous studies in congenitally athymic nude rats have suggested that the thymus is important for the development of intestinal T cells. Here we have examined the effect of the nude mutation on intraepithelial lymphocyte (IEL) development from the perinatal period. By immunohistochemistry it was shown that CD3−CD8αα+ putative IEL precursors colonized the epithelium of both normal and athymic neonatal rats. Mature T cells, however, did not develop in athymic neonates. In normal rats, γδ T cells were present at birth and αβ T cells appeared within 8 days of postnatal life. At this age, the composition and relative number of intraepithelial T cells were similar to that in normal adult rats, with the exception that most neonatal T‐cell receptor‐γδ+ and ‐αβ+ IEL expressed CD8β. By contrast, extrathymic T‐cell maturation in the gut of congenitally athymic rats occurred slowly, as CD3+ IEL did not appear until 4–6 months of age. These intraepithelial T cells displayed variable phenotypes and appeared to be induced by environmental antigens as they were not found in isolator‐kept old nudes. In conclusion, the present results indicate that the major colonization of the gut epithelium with γδ and αβ T cells expressing CD8αβ takes place perinatally and requires the presence of the thymus. The developmental relationship between these neonatal T cells and more immature CD3− CD8αα+/− IEL remains elusive.
Adenosine decreases post‐ischaemic cardiac TNF‐α production: anti‐inflammatory implications for preconditioning and transplantation
Authors: MELDRUM D. R. , CAIN B. S. , CLEVELAND JR. J. C. , MENG X. , AYALA A. , BANERJEE A. , HARKEN A. H.
Published: November 2003

Tumour necrosis factor‐α (TNF‐α) is an autocrine contributor to myocardial dysfunction and cardiomyocyte death in ischaemia–reperfusion injury (I/R), sepsis, chronic heart failure and cardiac allograft rejection. Cardiac resident macrophages, infiltrating leucocytes, and cardiomyocytes themselves produce TNF‐α. Although adenosine reduces macrophage TNF‐α production and protects myocardium against I/R, it remains unknown whether I/R induces an increase in cardiac TNF‐α in a crystalloid‐perfused model (in the absence of blood), and, whether adenosine decreases cardiac TNF‐α and protects function after I/R. To study this, isolated rat hearts were crystalloid‐perfused using the Langendorff method and subjected to I/R, with or without adenosine pretreatment. Post‐ischaemic cardiac TNF‐α (enzyme‐linked immunosorbent assay and bioassay) and function were determined (Langendorff). I/R increased cardiac TNF‐α and impaired myocardial function. Adenosine decreased cardiac TNF‐α and improved post‐ischaemic functional recovery. This study demonstrates that: first, I/R induces an increase in cardiac tissue TNF‐α in a crystalloid‐perfused model; second, adenosine decreases cardiac TNF‐α and improves post‐ischaemic myocardial function; third, decreased cardiac TNF‐α may represent a mechanism by which adenosine protects myocardium; and fourth, adenosine‐induced suppression of cardiac TNF‐α may provide an anti‐inflammatory link to preconditioning and have implications for cardiac allograft preservation.
The high‐affinity FcγRI on PMN: regulation of expression and signal transduction
Authors: HOFFMEYER F. , WITTE K. , SCHMIDT R. E.
Published: November 2003

In this paper we report that interferon‐γ(IFN‐γ) maintains and enhances functional properties of terminally differentiated polymorphonuclear cells (PMN). Such effects are obvious when compared with untreated PMN declining in their functional response. Culture for 18 hr with 100 IU/ml of recombinant IFN‐γ resulted in the expression of about 15 000 FcγRI per PMN. IFN‐γ maintained viability of PMN and prevented reduction of FcγRIIIb caused by apoptosis. No alterations were found in the level of FcγRIIa. Next, functional properties of FcγRI were evaluated. Calcium mobilization was detected in fura‐2/acetoxymethylester (AM)‐loaded PMN using a spectrophotometer and the respiratory burst was measured as luminol‐dependent chemiluminescence. Cross‐linking of an F(ab′)2 fragment of monoclonal antibody (mAb) 22.2 to FcγRI gave similar results to those obtained with intact monomeric human immunoglobulin G (mhIgG) when subsequently cross‐linked. Moreover, after blocking FcγRIIa and FcγRIIIb with respective mAb fragments, mhIgG was still effective in triggering a calcium flux demonstrating that second messenger generation was caused by FcγRI engagement. Even though FcγRI expression was lower than that of FcγRIIa, FcγRI induced a higher increase of the respiratory burst. In addition, the protein tyrosine phosphatase CD45 was able to inhibit both responses when it was co‐cross‐linked with CD64, suggesting involvement of tyrosine phosphorylation in early signalling steps.
T‐cell‐independent granuloma formation in response to Mycobacterium avium: role of tumour necrosis factor‐α and interferon‐γ
Authors: SMITH D. , HÄNSCH H. , BANCROFT G. , EHLERS S.
Published: November 2003

SUMMARY We used Mycobacterium avium infection in severe combined immunodeficiency (SCID) mice to examine T‐cell‐independent mechanisms of inflammatory cell recruitment. SCID mice infected with a virulent strain of M. avium (TMC724) were able to recruit macrophages to sites of mycobacterial replication and formed organized and coherent granulomas in the absence of functional T cells. Phagocyte recruitment was almost totally ablated by neutralization of either tumour necrosis factor‐α (TNF‐α) or interferon‐γ (IFN‐γ) in vivo demonstrating that granuloma formation was dependent on the presence of these cytokines. This was concomitant with a reduction in the in situ cytokine mRNA levels otherwise induced in infected mice, for chemokines, pro‐inflammatory and regulatory cytokines, including TNF‐α, IFN‐γ, macrophage inflammatory protein‐1α, interleukin‐1β (IL‐1β) and IL‐10. Furthermore, in vivo treatment of infected mice with anti‐asialo GM‐1 antisera, which depletes natural killer (NK) cells, prevented recruitment of inflammatory cells. In vitro studies confirmed that M. avium was able to elicit IFN‐γ from SCID spleen in a dose‐dependent manner. These data show for the first time that secretion of IFN‐γ from NK cells can mediate a T‐cell‐independent pathway of granuloma formation and cellular infiltration in response to mycobacteria.
The major dog allergens, Can f 1 and Can f 2, are salivary lipocalin proteins: cloning and immunological characterization of the recombinant forms
Authors: KONIECZNY† A. , MORGENSTERN‡ J. P. , BIZINKAUSKAS C. B. , LILLEY C. H. , BRAUER§ A. W. , BOND J. F. , AALBERSE R. C. , WALLNER** B. P. , KASAIAN M. T.
Published: November 2003

Canis familiaris allergen 1 (Can f 1) and Canis familiaris allergen 2 (Can f 2) are the two major allergens present in dog dander extracts. We now report the isolation of cDNAs encoding both proteins and present their nucleotide and deduced amino acid sequences. Can f 1, produced by tongue epithelial tissue, has homology with the von Ebner’s gland (VEG) protein, a salivary protein not previously thought to have allergenic properties. Can f 2, produced by tongue and parotid gland, has homology with mouse urinary protein (MUP), a known allergen. Both VEG protein and MUP are members of the lipocalin family of small ligand‐binding proteins. Recombinant forms of Can f 1 and Can f 2 were produced and tested for immunoglobulin E (IgE) reactivity. Among dog‐allergic subjects, 45% had IgE directed exclusively to rCan f 1, and 25% had IgE to both rCan f 1 and rCan f 2. In addition, both recombinant proteins were able to cross‐link IgE and elicit histamine release from peripheral blood leucocytes in vitro. These findings confirm that Can f 1 and Can f 2 are major and minor dog allergens, respectively, and demonstrate that recombinant forms of dog allergens retain at least some IgE‐binding epitopes.
Glucocorticoids enhance concanavalin A‐induced mitogenic response through the inhibition of nitric oxide production
Authors: RAMÍREZ F. , SILVA A.
Published: November 2003

Glucocorticoids (GC) are known to inhibit mitogen‐induced proliferation of T cells. In this study we show two experimental situations where the addition of GC increases lymphocyte proliferation. It has been reported by different authors that rat spleen (SPL) cells proliferate poorly after concanavalin A (Con A) activation. These poor responses have been related to the suppressor activity of macrophages. Similarly, it is known that T‐cell proliferation is depressed in the presence of an excess of macrophages in the culture. Here we show that in both experimental situations, the inclusion of dexamethasone (DEX), a synthetic glucocorticoid, in the culture medium enhances the Con A‐stimulated proliferation. We provide evidence that this effect is a consequence of the inhibition of nitric oxide (NO) synthesis by the hormone. Furthermore, we also demonstrate that rat SPL cells are inefficient antigen‐presenting cells (APC) because of their spontaneous high production of NO. Taken together our results suggest that the effects of GC on T‐cell activation may be to promote or inhibit proliferation depending on the level of endogenous NO synthesis. The possible significance of these results is briefly discussed.
Characterization of a constitutive type III nitric oxide synthase in human U937 monocytic cells: stimulation by soluble CD23
Authors: ROMAN V. , DUGAS N. , ABADIE A. , AMIRAND C. , ZHAO H. , DUGAS B. , KOLB J.‐P.
Published: November 2003

The soluble cleavage fragment of the low‐affinity immunoglobulin E (IgE) receptor/CD23 (sCD23 25 000 MW) and antibodies directed against their receptors on monocytes, CD11b and CD11c, stimulate the production of nitric oxide (NO) by these cells and we have suggested that the enzyme involved could be related to the endothelial constitutive type III nitric oxide synthase (ecNOS). In the present work, we have analysed the characteristic properties of this NOS isoform in the model of the human promonocytic cells U937 By reverse‐transcription polymerase chain reaction (RT‐PCR), the presence of an mRNA coding for type III NOS was found in U937 cells and the corresponding protein was detected by immunofluorescence in permeabilized cells with a specific anti‐ecNOS monoclonal antibody (mAb). Membrane extracts displayed a NOS activity dependent on the presence of calcium and calmodulin in the reaction medium and that was abrogated in the presence of EGTA. Recombinant soluble CD23 (25 000 MW) was found to trigger an NO‐dependent cGMP accumulation in these cells, which was abrogated by calcium chelators and inhibitors of the calcium/calmodulin complex. Moreover, sCD23 elicited a transient augmentation of intracytoplasmic free calcium concentration [ Ca2+]i that was dependent on the presence of calcium in the external buffer and was prevented in the presence of EGTA, indicating that it was due to a calcium influx. In conclusion, human promonocytic cells such as U937 exhibit a functional type III NOS that can be stimulated by calcium‐raising agents, such as sCD23
TAK‐603 selectively suppresses Th1‐type cytokine production and inhibits the progression of adjuvant arthritis
Authors: OHTA Y. , YAMANE M. , SOHDA T. , MAKINO H.
Published: November 2003

We have shown that TAK‐603, a new anti‐rheumatic drug, is more effective in animal models in which cellular immunity plays a central role. Here, we studied the effect of the drug on Th1 cytokines, which are dominantly produced in this type of immune reaction, in an in vitro system and an in vivo model. We established Th1‐ and Th2‐dominant T‐cell lines, and studied the effect of TAK‐603 on their cytokine production. Th1 cell lines were BALB/c mouse allo‐reactive T cells and C57BL mouse mite antigen‐reactive T cells, and the Th2 cell line was BALB/c mouse ovalbumin‐reactive T cells. TAK‐603 suppressed the production of Th1 cytokines [
interferon‐&ggr; (IFN‐&ggr;) and interleukin‐2 (IL‐2)] and not that of Th2 cytokines (IL‐4, IL‐5) in these cell lines. Furthermore, selective suppression of Th1 cytokine production was also observed in the T‐cell clones obtained from the ovalbumin‐reactive T‐cell line. To investigate the effect on cytokine production in animal models of arthritis, we analysed the expression of cytokine messenger RNA using reverse transcription–polymerase chain reaction. In adjuvant arthritis rats, Th1‐dominant cytokine production was observed both in the arthritic joint and the spleen, and the time–course paralleled the progression of arthritis. On the other hand, in type‐II collagen‐induced arthritis, in which TAK‐603 has little effect, Th1‐dominant cytokine production was not observed and Th2 cytokines were shown to be more important. The adjuvant arthritis rats treated with TAK‐603 (6·25 mg/kg/day, per os) showed significantly lower cytokine mRNA expression both locally and systemically. These data suggest that TAK‐603 selectively suppresses Th1 cytokine production, which is consistent with its effect on cellular immunity in animal models.
Mobile information systems — executives' view
Authors: Puuronen S. , Savolainen V.
Published: November 2003

The concepts of travelling executive and executives' mobile information system are first defined in this paper. The main findings are in the form of collected data and opinions concluded from our personal discussions with 49 executives in the United Kingdom, France, Italy and Finland concerning the nature of the work of executives and the usage of information technology (IT) to support their work today. The near future expectations of the executives are also analysed, especially concerning the mobile use of IT services in order to construct executives' holistic view of mobile computing. The use of IT services was found to be very widespread. Big differences were found: some of the executives had clear positive attitudes for using several mobile IT services but many had not yet realized the potential advantages of these services. Many reasons, e.g. certain cultural differences, were found to explain differences in these four countries.
Studying system development methodologies: an examination of research methods
Authors: Wynekoop Judy L. , Russo Nancy L.
Published: November 2003

Changes in organizational structures, competitive environments, technologies and economies are altering the types and complexity of computer‐based information systems. For methodologies, tools and techniques to evolve to meet these changing needs, research to assess current system development practices would be expected. However, there is little evidence that the evaluation of methodologies and improvement of the systems development process are keeping pace with technological and organizational changes. This paper analyses existing research on system development methodologies and discusses the research needed to provide the information scholars and practitioners will need for the evaluation, selection and development of methodologies in a changing environment.
The use of RepGrids to gather interview data about information systems analysts
Authors: Hunter M. Gordon
Published: November 2003

In view of suggestions for the application of more qualitative and interpretive approaches to information systems research, this article describes the use of RepGrids to gather interview data about information systems analysts. In the light of the objectives of a specific research project, various techniques are considered. Analysis of the techniques in relation to specific research project objectives leads to the adoption of a form of Kelly's RepGrid technique for the specific project described here. A detailed description is included about how RepGrids were applied in this instance. It is shown that RepGrids are appropriate for gathering qualitative interview data. The technique provides structure to a ‘one‐to‐one’ interview. But, at the same time, RepGrids allow sufficient flexibility for the research participants to be able to express their own interpretation about a particular topic. Further, because the content of the data is determined by the research participant researcher bias is reduced. The article concludes by suggesting that the RepGrid technique supports the generation of rich in‐depth data and is a reliable research technique for gathering qualitative data about information systems analysts.
The rise and fall of an executive information system: a case study
Authors: McBride Neil
Published: November 2003

The progress of an executive information system project within a manufacturing organization over a period of 9 years is described. The case study illustrates the importance of the interaction between the business environment, the organizational environment and the perceptions and interpretations of events and facts by stakeholders on the success or failure of an information system. It shows the importance of context in the development and implementation of an executive information system and the dynamic nature of the influence of social, economic and technical factors. The reasons for the initial success and the subsequent failure of the EIS within the company are explored from a contingency perspective.
The use of systems development methodologies in practice: a field study
Authors: Fitzgerald B.
Published: November 2003

This paper discusses the findings of a field study of experienced systems developers which investigated the use of systems development methodologies (SDMs) in practice. Among the main findings to emerge are the following. There is a wide difference between the formalized sequence of steps and stages prescribed by a methodology and the methodology‐in‐action uniquely enacted for each development project. Also, there is much evidence which suggests that developers omit certain aspects of methodologies not from a position of ignorance, but from the more pragmatic basis that certain aspects are not relevant to the development environment they face. Thus, methodologies are tailored to the needs of the prevailing development environment and, being documented in a single manual often, are framed at a higher level of granularity that specifies broad activities and objectives, rather than specifying in a very detailed fashion the exact manner in which development should take place. Additionally, the findings clarify the apparently contradictory position in the literature as to whether developer experience is positively or negatively correlated with methodology use. The study would suggest that experienced developers are likely to use methodologies, albeit ones which have been heavily customized to the exigencies of the particular situation.
Regulation of the ecdysone receptor, USP, E75 and MHR3 mRNAs by 20‐hydroxyecdysone in the GV1 cell line of the tobacco hornworm,Manduca sexta
Authors: Lan Q. , Wu Z. , Riddiford L. M.
Published: November 2003

The responsiveness of several nuclear transcription factor genes to 20‐hydroxyecdysone (20E) was characterized in an embryonic cell line, GV1, from

Manduca sexta

. The mRNA for the

Manduca

ecdysone receptor (

Ms

EcR) was present in the GV1 cells and transiently increased 2.3‐fold by 5 h after the addition of 2 μg/ml (4 × 10−6
M
) 20‐hydroxyecdysone (20E). In contrast,

Manduca

ultraspiracle (

Ms

USP) mRNA level in the GV1 cells decreased slowly to half of its initial level by 12 h when exposed to the same concentration of 20E. The mRNAs for two putative transcription factors,

Ms

E75 and MHR3, were induced in the GV1 cells by 20E, with that for E75 appearing within 1 h whereas that for MHR3 within 2 h. The ED50s for induction of

MsE75

and

MHR3

gene expression in GV1 cells were 1.2 × 10−6 and 2.5 × 10−6
M
20E, respectively.
Identification and mRNA developmental profiles of two ultraspiracle isoforms in the epidermis and wings ofManduca sexta
Authors: Jindra M. , Huang J‐Y. , Malone F. , Asahina M. , Riddiford L. M.
Published: November 2003

cDNAs were isolated from

Manduca sexta

that encode two isoforms of an ultraspiracle (USP) homologue MsUSP‐1 and MsUSP‐2 with different N‐terminal A/B regions. The MsUSP‐1 cDNA predicts a protein with 97% and 45% amino acid identities in the DNA‐ and ligand‐binding domains respectively to the

Drosophila

USP and 89% overall identity with

Bombyx mori

CF1 (an USP homologue). Northern blot hybridizations with probes specific to MsUSP‐1 and MsUSP‐2 showed transcripts of an approximately equal size (4.5 kb), but with diverse developmental profiles in

Manduca

epidermis during the two final larval instars and the onset of the adult moult. The MsUSP‐1 mRNA was expressed during the intermoult periods, with higher levels around the time of the larval ecdyses and at the onset of wandering behaviour. In contrast, the MsUSP‐2 mRNA was up‐regulated at times of high ecdysteroid titre during the larval moults, when the MsUSP‐1 mRNA disappeared. Together, these conversely regulated isoform mRNAs contribute to the constitutive expression profile of total MsUSP mRNA.
Juvenile hormone controls early trypsin gene transcription in the midgut ofAedes aegypti
Authors: Noriega F. G. , Shah D. K. , Wells M. A.
Published: November 2003

Early trypsin is a female‐specific protease present in the

Aedes aegypti

midgut during the first few hours after ingestion of a blood meal. The enzymatic activity of early trypsin plays an essential role in the transcriptional activation of the late trypsin gene, which encodes the major midgut endoprotease involved in blood meal protein digestion. Transcription of the early trypsin gene is part of the normal post‐emergence maturation of the midgut in the adult female. Abdominal ligation within 1 h of emergence completely prevented the transcription of the early trypsin gene. Topically applied JH III or methoprene induced transcription of the early trypsin gene in ligated abdomens to levels similar to those observed in non‐ligated females. The induction of early trypsin transcription by JH is dose‐dependent and ‘head‐independent’, suggesting that factors coming from the neurosecretory axis are not required.
Phenobarbital induction of CYP6D1 is due to atransacting factor on autosome 2 in house flies,Musca domestica
Authors: Liu N. , Scott J. G.
Published: November 2003

To improve our understanding of phenobarbital (PB) mediated induction of CYP6D1 we examined the genetic linkage of PB induction using PB responsive (aabys) and non‐responsive (LPR) strains of house flies. PB induction was linked to autosome 2, indicating that

CYP6D1

is

trans

regulated by this PB inducible element. Our results are discussed relative to a previous hypothesis that the same regulatory genes are responsible for both induction and monooxygenase‐mediated insecticide resistance.
Mitochondrial haplotypes and phylogeography of Phlebotomus vectors of Leishmania major
Authors: Esseghir S. , Ready P. D. , Killick‐Kendrick R. , Ben‐Ismail R.
Published: November 2003

Haplotypes of eight phlebotomine species were characterized by cycle sequencing a mitochondrial (mt) DNA fragment (cytochrome b to NADH1) amplified from single sandflies by PCR. Phlebotomus (Phlebotomus) papatasi displayed little variation throughout its large geographical range. We conclude that this vector of Leishmania major suffered a population bottleneck late in the Pleistocene and then radiated out from the eastern Mediterranean subregion. There was no support for a recent domestic lineage of P. papatasi. The mtDNA molecular clock in phlebotomines (subgenera Phlebotomus and Larroussius) was calibrated by reference to palaeogeographical events in Africa and the Mediterranean subregion. It fitted a pairwise nucleotide sequence divergence rate of 1.0–2.5% per million years. Co‐evolution of L. major, its Phlebotomus vectors and mammalian reservoirs is discussed.
Reassessment of 79B actin gene expression in the abdomen of adult Drosophila melanogaster
Authors: Ohshima S. , Villarimo C. , Gailey D. A.
Published: November 2003

The expression pattern of the 79B actin gene in Drosophila melanogaster has been inferred previously by means of a reporter gene in which 79B actin promoter sequences drive the lacZ coding sequences. Although the 79B actin gene is expressed primarily in muscles of the thorax and first abdominal segment of the adults of both sexes, expression in the remaining abdominal segments appears limited to the male genital muscles and the male‐specific Muscle of Lawrence (MOL). This reported abdominal expression pattern has been reassessed. By varying parameters of tissue preparation and lacZ reporter gene detection, expression of the 79B actin gene has been revealed in most dorsal abdominal longitudinal fibres and genital muscles of both females and males. These new results suggest that there are differences in the level of 79B actin gene expression among the various abdominal muscles of both sexes, and that abdominal expression is not limited primarily to male sex structures.
The phylogenetic relationshipsof seven European Ips (Scolytidae, Ipinae) species
Authors: Stauffer C. , Lakatos F. , Hewitt G. M.
Published: November 2003

In Central Europe seven Ips species are characterized by differences in morphology, structure of galleries, host specificity and aggressiveness. These species were analysed by allozyme markers and by sequencing 567 bp of the mitochondrial cytochrome oxidase I gene, in order to define their phylogenetic relationships. Orthotomicus erosus and Tomicus minor were taken as outgroup species. The data revealed high inter‐specific and low intra‐specific sequence divergence. Plotting the observed sequence divergence caused by transversions (Tv) and transitions (Ts) and the level of saturation for Ts and Tv of each codon position showed that the third positions were highly saturated by multiple substitutions. Maximum parsimony analysis produced two groups: (1) I. typographus, I. cembrae, I. amitinus, I. duplicatus and I. acuminatus; (2) I. mannsfeldi, I. sexdentatus and the two outgroups. In all analyses the species of the first cluster were put together and I. typographus and I. cembrae, and I. mannsfeldi and O. erosus emerged as sister pairs.

The data do not support a common ancestor for the seven European Ips species. The eight‐spined bark beetles (except I. mannsfeldi) and I. acuminatus formed a monophyletic group. The close relationship of I. mannsfeldi and O. erosus supports the latter belonging to the genus Ips as proposed by Wood (1982) and Escherich (1923). However, more genetic markers and more species of the genera Orthotomicus and Pityokteines have to be analysed to resolve the phylogenetic positions of I. sexdentatus and I. mannsfeldi within the tribe Ipini.
Mosquito clathrin heavy chain: analysis of protein structure and developmental expression in the ovary during vitellogenesis
Authors: Kokoza V. A. , Snigirevskaya E. S. , Raikhel A. S.
Published: November 2003

We have deduced the amino acid sequences of clathrin heavy chain (CHC) polypeptides based on cDNA and genomic clones from the mosquito, Aedes aegypti. Two isoforms which differ in the very beginning of the N‐terminal domain, ovary‐specific AaCHCa and somatic‐specific AaCHCb, were identified, characterized and compared to one another as well as to CHC polypeptides from different species. The 1682 amino acid sequence of the AaCHCa isoform predicts a molecular mass (Mr) of 191,743 daltons and an isoelectric point of 5.80, whereas the 1674 amino acid sequence of the AaCHCb isoform predicts a Mr of 191,033 daltons and an isoelectric point of 5.71. Both mosquito AaCHC isoforms are highly conserved, with full‐sequence identities of 88% to Drosophila melanogaster, 81% to mammal (rat, cow and human), 71% to C. elegans, 58% to Dictyostelium discoideum, and 49% to yeast CHC polypeptides. The highest degree of conservation is in the middle portion of the mosquito CHC molecule which includes the linker region and extended triskelion arm, with decreasing conservation through the N‐terminal domain, trimerization domain, and the relatively diverged C‐terminal region. The protein domains do not directly correspond to specific exons of the mosquito AaCHC gene, with the exception of exon 6 which encodes the C‐terminal domain of the CHC polypeptide. Polyclonal antibodies raised against a bacteria‐expressed AaCHC fusion protein recognized one major band of about 180 kDa in vitellogenic ovary whole‐lysate. Immunogold labelling of the AaCHC polypeptide localized it to the coat of coated pits and coated vesicles in oocytes from vitellogenic follicles. Northern blot and in situ hybridization analyses suggest that regulation of AaCHC gene expression in the ovary is complex, and it likely involves both developmental and hormonal signals.
An in‐vitro evaluation of sealer placement methods
Authors: KAHN F. H. , ROSENBERG P. A. , SCHERTZER L. , KORTHALS G. , NGUYEN P. N. T.
Published: November 2003

Successful root canal therapy is dependent upon obturating the root canal system. A critical component of the filling procedure is sealer placement. This in vitro study investigated the efficacy of six methods of sealer placement using clear plastic blocks with simulated curved canals instrumented to the apex using a stepback technique. After sealer placement and setting, the blocks were cross‐sectioned at 2 mm intervals from midcanal to apex. The sections were evaluated with a stereo operating microscope at × 6 magnification for placement of an evently distibuted layer of sealer around the prepared canal walls. The lentulo spiral and the Max‐i‐Probe Delivery System were the most effective means of sealer placement, followed by ultrasonic and sonic files. The least effective methods were the paper point and the K file.
Preparation of simulated root canals using the Macfile, Canal Master U and K‐FIexofile
Authors: MARTIN G. , BLAŠKOVIćG‐ŠUBAT V.
Published: November 2003

The efficacy of three different root canal instruments in maintaining the original shape of curved canals was evaluated. Acrylic blocks were prepared to simulatecanals with 40° and 60° curvature. K‐Flexofiles and Macflles were used with a step‐back technique utilizing an ‘in/out’ filing motion. The balanced‐force technique was used with Canal Master ‘U’ instruments. Ten 40° and 10 60° canals were prepared with each instrument.The results were evaluated using superimposed photo‐graphic slides at X20 magnification to show the canals before and after the instrumentation. The width of the prepared canals was measured at their convex/concave point to determine the divergence of the instrumented canal shape from the original canal shape. The measure‐ments were taken at the level of the apical foramen and 3 and 6 mm coronal to it. In this way it was possible to recognize all the defects created during the preparation of a canal. Among the tested instruments, the Macfile provoked minimal canal deviation (P=0.05).The Canal Master ‘U’ was similarly effective (P=0.05), but had the highest incidence of instrument fracture. The K‐Flexofile demonstrated the most defects in canal preparation and canal deviation, particularly at the level of the apical foramen (P< 0.01)
Thermographic assessment of heat generated on the root surface during post space preparation
Authors: HUSSEY D. L. , BIAGIONI P. A. , MCCULLAGH J. J. P. , LAMEY P.‐J.
Published: November 2003

The mechanical preparation of a post space in a root filled tooth is commonly carried out using rotating instruments in a slow handpiece. This procedure creates frictional heat which may be transferred to the root surface. In this investigation, the temperature changes on the root surface of 12 root filled, extracted human premolar teeth during preparation of post spaces were analysed using an infrared thermal imaging camera. A rotational speed of 8000 r.p.m. was used and the teeth were prepared with size 2 and 3 Peeso reamers and a red Parapost drill to the optimal length. The results indicate that very large temperatures are generated on the root surface and that this is greatest during the Parapost preparation stage. The long‐term clinical effects of post space preparation by engine driven drills would warrant further investigation.
Thermographic assessment of root canal obturation using thermomechanical compaction
Authors: MCCULLAGH J. J. P. , BIAGIONI P. A. , LAMEY P.‐J. , HUSSEY D. L.
Published: November 2003

The temperature changes on the root surface of 30 extracted human premolar teeth during thermomechanical root canal obturation with gutta‐percha were determined using an infrared thermal imaging camera. Three handpiece rotational speeds of 8, 12 and 16 × 103 r.p.m. were used, in conjunction with a Gutta Condensor. On completion of the procedure, the quality of tooth canal obturation was examined radiographically. Under the conditions of this experiment, surface root temperature rises of >97° were recorded during all three speed settings. The radiographic quality of obturation between the groups appeared not to be significantly different. The clinical relevance of these findings is uncertain, but the temperatures reported are of a magnitude to be of biological importance.
Short‐term tissue response to potential root‐end filling materials in infected root canals
Authors: CHONG B. S. , PITT FORD T. R. , KARIYAWASAM S. P.
Published: November 2003

The short‐term tissue responses to two potential root‐end filling materials, a light‐cured glass ionomer cement (Vitrebond) and a reinforced zinc oxide–eugenol cement (Kalzinol), were compared with that to amalgam using a previously devised experimental model. In 24 premolar teeth of beagle dogs (47 roots), a collection of endodontic pathogenic bacteria was first inoculated into the root canals to induce periradicular lesions. On each root, an apiccctomy was performed and root‐end cavities prepared to receive fillings of each material. The teeth and surrounding jaw were removed after 2 weeks (23 roots) and 1 week (24 roots); they were then prepared for histological examination. The tissue response to amalgam fillings after 2 weeks and 1 week was marked by moderate or severe inflammation on all roots, and extended to 0.5 mm or >0.5 mm in 15 out of 16 roots. In contrast, after 2 weeks, the majority of roots filled with Kalzinol showed little or moderate inflammation, while the tissue response to Vitrebond was the best of the three materials, and was also the least extensive. After 1 week, the overall best tissue response was with Vitrebond, followed by Kalzinol. The differences between materials for both time periods with either none or few inflammatory cells when compared with that with either moderate or severe inflammation were not statistically significant (P<0.02). However, the differences between materials for both time periods with no inflammation or inflammation extending <0.2 mm when compared with that with inflammation extending >0.2 mm (0.5 mm or >0.5 mm) were statistically significant (P<0.01). Apart from amalgam, in which healing was marked by the persistence of a localized focus of inflammation adjacent to the root‐end filling, even though there were intersamlpe variations, there was little overall difference in the temporal and qualitative healing response to Vitrebond and Kalzinol. Both Vitrebond and Kalzinol have potential as root‐end filling materials, as the tissue response was considerably more favourable than that to amalgam even in the short‐term.