bluuebunny/crossref_metadata_2025_split huggingface dataset:
The contribution of both oxygen and nitrogen intermediates to the intracellular killing mechanisms of C1q‐opsonized Listeria monocytogenes by the macrophage‐like IC‐21 cell line
Authors: Álvarez‐Domínguez C. , Carrasco‐Marín E. , López‐Mato P. , Leyva‐Cobián F. 
Published: March 2003

Summary Listeria monocytogenes is a facultative intracellular pathogen which is internalized by host mammalian cells upon binding to their surface. Further listerial growth occurs in the cytosol after escape from the phagosomal–endosomal compartment. We have previously reported that C1q is able to potentiate L. monocytogenes phagocytosis upon bacterial opsonization by ingestion through C1q‐binding structures. In this report, we analysed the post‐phagocytic events upon internalization of C1q‐opsonized L. monocytogenes and found an induction of macrophage (Mφ)‐like IC‐21 cell bactericidal mechanisms displayed by the production of oxygen and nitrogen metabolites. Both types of molecules are effective in L. monocytogenes killing. Further analysis of the cellular responses promoted by interaction of C1q with its surface binding structures, leads us to consider C1q as a collaborative molecule involved in Mφ activation. Upon interaction with surface binding structures, C1q was able to trigger and/or amplify the production of reactive oxygen and nitrogen intermediates induced by stimuli such as interferon‐γ and L. monocytogenes phagocytosis.
M‐ficolin is expressed on monocytes and is a lectin binding to N‐acetyl‐	d‐glucosamine and mediates monocyte adhesion and phagocytosis of Escherichia coli
Authors: Teh C. , Le Y. , Lee S. H. , Lu J. 
Published: March 2003

SummaryFicolins are a group of multimeric proteins that contain collagen‐like and fibrinogen‐like (FBG) sequences. Three types of ficolins have been characterized: H‐, L‐ and M‐ficolins. Both H‐ and L‐ficolins have demonstrated lectin activities. In the present study, the FBG domain of M‐ficolin was expressed and shown to bind to N‐acetyl‐
d‐glucosamine. M‐ficolin mRNA was expressed in monocytes but not in the more differentiated macrophages and dendritic cells. By flow cytometry, surface biotinylation and immunoprecipitation, we showed that M‐ficolin was associated with the surface of promonocytic U937 cells. M‐ficolin transiently expressed in COS‐7 cells was also clearly detected on the cell surface by immunoprecipitation. By flow cytometry, M‐ficolin was detected on peripheral blood monocytes but not on lymphocytes or granulocytes. Immobilized rabbit anti‐M‐ficolin F(ab′)2 mediated U937 cell adhesion, and the antibody also inhibited phagocytosis of Escherichia coli K‐12 by U937 cells. Therefore, M‐ficolin might act as a phagocytic receptor or adaptor on circulating monocytes for micro‐organism recognition and may potentially mediate monocyte adhesion.
Modulation of CD11C+ splenic dendritic cell functions in murine visceral leishmaniasis: correlation with parasite replication in the spleen
Authors: Basu A. , Chakrabarti G. , Saha A. , Bandyopadhyay S. 
Published: March 2003

SummaryBALB/c mice resolve Leishmania donovani infection in the liver over an 8–12‐week period. However, after an initial phase of 2–4 weeks where increases in parasite load are not readily detectable, parasite numbers in the spleen begin to increase reaching maximum levels at 16 weeks post‐infection. Thereafter, parasite replication in the spleen is controlled and BALB/c mice maintain this residual parasite load in the spleen for many months, without further increase. We evaluated functions of CD11C+ splenic dendritic cells throughout the course of L. donovani infection in the spleen of BALB/c mice. Unlike the dendritic cell (DC)‐specific antigen DEC‐205, CD11C was not up‐regulated on macrophages during visceral leishmaniasis. No appreciable impairment of splenic DC functions was observed when this antigen‐presenting cell subset was purified from 30‐day post‐infected mice. Significant impairment in inducing allogeneic mixed lymphocyte reaction (MLR) and presenting L. donovani antigens or keyhole limpet haemocyanin (KLH) to specific T cells was observed with CD11C+ splenic DC purified from 60‐day post‐infected mice. Functional impairment of splenic DC at 60 days post‐infection correlated with their reduced surface expression of major histocompatibility complex (MHC) class II molecules, impairment of interleukin‐12 (IL‐12) production and to their ability to suppress interferon‐γ (IFN‐γ) production by Leishmania antigen‐primed T cells. Of interest, the impairment of splenic DC in presenting Leishmania antigens or KLH to specific T cells was corrected at 120 days post‐infection, and correlated with their up‐regulation of MHC class II expression, IL‐12 production, induction of IFN‐γ by Leishmania antigen‐primed T cells and the onset of control over splenic parasite replication in vivo. These results indicate that functional integrity of DC may be important in controlling L. donovani infection.
Production and functional analysis of rat CD59 and chimeric CD59‐Crry as active soluble proteins in Pichia pastoris
Authors: Quigg R. J. , He C. , Hack B. K. , Alexander J. J. , Morgan B. P. 
Published: March 2003

SummaryCrry (CR1‐related gene/protein) is a rodent complement regulator that inhibits C3 convertases. CD59 is a conserved protein inhibitor active towards C8 and C9. We have previously produced rat Crry as a recombinant soluble (rs) protein in Pichia pastoris. In this study we produced functionally active rat rsCD59 and a chimeric rsCD59‐Crry protein in P. pastoris. The GPI anchor addition site of rat CD59 (Asn‐79) was replaced either by a stop codon to produce rsCD59, or with the sequence of the first five short consensus repeats of Crry to produce rsCD59‐Crry. Proteins were generated by fermentation and purified by affinity chromatography on an anti‐CD59 column. In a standard classical pathway haemolysis assay, all three rs proteins had inhibitory activity, with 50% inhibition at 0·5 µm (rsCrry and rsCD59‐Crry) and 4·4 µm (rsCD59). In an assay examining inhibition of C5b‐9, in which C5b‐7 was first formed, followed by purified C8 and C9, rsCD59 and rsCD59‐Crry were active with 50% inhibition at 0·8 µm (rsCD59‐Crry) and 1·3 µm (rsCD59). The degree of inhibition was independent of whether the C8 and C9 were of rat or human origin. Therefore, we have produced rsCD59 and rsCD59‐Crry in P. pastoris. The rsCD59 retains its inhibitory activity towards C5b‐9, while rsCD59‐Crry appears to have the combined activities of Crry and CD59. In a haemolytic assay, the inclusion of CD59 to Crry is of no additional benefit to Crry, which may illustrate the overall importance of the C3 convertase step. Yet, inclusion of Crry to CD59 increases the potency of CD59 towards C5b‐9.
CXC chemokine receptor 4 expression and stromal cell‐derived factor‐1α‐induced chemotaxis in CD4+ T lymphocytes are regulated by interleukin‐4 and interleukin‐10
Authors: Jinquan T. , Quan S. , Jacobi H. H. , Madsen H. O. , Glue C. , Skov P. S. , Malling H.‐J. , Poulsen L. K. 
Published: March 2003

SummaryWe report that interleukin (IL)‐4 and IL‐10 can significantly up‐ or down‐regulate CXC chemokine receptor 4 (CXCR4) expression on CD4+ T lymphocytes, respectively. Stromal cell‐derived factor‐1α (SDF‐1α)‐induced CD4+ T‐lymphocyte chemotaxis was also correspondingly regulated by IL‐4 and IL‐10. IL‐4 and IL‐10 up‐ or down‐regulated CXCR4 mRNA expression in CD4+ T lymphocytes, respectively, as detected by real‐time quantitative reverse transcription–polymerase chain reaction (RT–PCR). Scatchard analysis revealed a type of CXCR4 with affinity (Kd≈ 6·3 n
m), and ≈ 70 000 SDF‐1α‐binding sites per cell, among freshly isolated CD4+ T lymphocytes, and two types of CXCR4 with different affinities (Kd1≈ 4·4 n
m and Kd2≈ 14·6 n
m), and a total of ≈ 130 000 SDF‐1α‐binding sites per cell, among IL‐4‐stimulated CD4+ T lymphocytes. The regulation of CXCR4 expression in CD4+ T lymphocytes by IL‐4 and IL‐10 could be blocked by a selective inhibitor of protein kinase (staurosporine) or by a selective inhibitor of cAMP‐ and cGMP‐dependent protein kinase (H‐8), indicating that these cytokines regulate CXCR4 on CD4+ T lymphocytes via both cAMP and cGMP signalling pathways. The fact that cyclosporin A or ionomycin were able to independently change the CXCR4 expression and block the effects of IL‐4 and IL‐10 on CXCR4 expression implied that the capacity of IL‐4 and IL‐10 to regulate CXCR4 on CD4+ T lymphocytes is not linked to calcium‐mobilization stimulation. These results indicate that the effects of IL‐4 and IL‐10 on the CXCR4–SDF‐1 receptor–ligand pair may be of particular importance in the cytokine/chemokine environment concerning the inflammatory processes and in the progression of human immunodeficiency virus (HIV) infection.
Mycobacterium tuberculosis‐activated dendritic cells induce protective immunityin mice
Authors: Tascon R. E. , Soares C. S. , Ragno S. , Stavropoulos E. , Hirst E. M. A. , Colston M. J. 
Published: March 2003

SummaryActivated dendritic cells are critically important in the priming of T‐cell responses. In this report we show that the infection of a conditionally immortalized dendritic cell line (tsDC) with Mycobacterium tuberculosis resulted in the up‐regulation of B7‐1 and B7‐2 co‐stimulatory molecules and the induction of several inflammatory cytokines, including tumour necrosis factor‐α and interleukin‐6, ‐1β and ‐12. In addition, we show that these activated dendritic cells were capable of eliciting antigen‐specific T‐cell responses and potent anti‐mycobacterial protective immunity in a murine model of experimental tuberculosis infection.
Demonstration that platelet‐activating factor is capable of activating mast cells and inducing a chemotactic response
Authors: Nilsson G. , Metcalfe D. D. , Taub D. D. 
Published: March 2003

SummaryPlatelet‐activating factor (PAF) is generated in a variety of inflammatory conditions in which mast cells accumulate. However, little is known about the ability of PAF to influence mast cell function directly. In this study we examine the ability of PAF to activate mast cells and regulate mast cell chemotaxis. PAF was found to induce intracellular calcium mobilization and chemotactic responses in both murine and human mast cells. PAF induced transient increases in intracellular Ca2+ concentrations with a 50% effective dose of 1 n
m and induced significant migratory responses at PAF concentrations of 1 n
m to 1 µm in the human leukaemia mast cell line (HMC‐1). Using signal transduction inhibitors, both PAF‐induced calcium mobilization and migration of mast cells were shown to require activation of pertussis toxin‐sensitive G proteins. PAF‐induced calcium and chemotactic responses were cross‐desensitized by C5a. Together, these data demonstrate that PAF is capable of activating distinct signalling pathways in mast cells associated with calcium mobilization and cell migration; and that PAF may thus contribute to the regulation of mast cell responses and hyperplasia at sites of inflammation.
The effects of DNA containing CpG motif on dendritic cells
Authors: Behboudi S. , Chao D. , Klenerman P. , Austyn J. 
Published: March 2003

SummaryDendritic cells (DC) are specialized antigen‐presenting cells. DC can acquire and process antigens in the periphery before maturing and migrating to secondary lymphoid tissues where they present the antigens and deliver co‐stimulatory signals to T cells. We describe an immunostimulatory oligonucleotide containing a CpG motif that stimulated murine DC to up‐regulate co‐stimulatory molecules, induce T‐cell proliferative responses and secrete interleukin‐12 in vitro. Administration of this oligonucleotide, but not of a control oligonucleotide lacking this motif, to mice led to the disappearance of DC from the marginal zone and T‐cell areas of spleen, but not from heart or kidney. The same CpG did not cause maturation of monocyte‐derived human DC in vitro, but lipopolysaccharide‐treated monocyte‐derived DC showed enhanced functional activity and up‐regulated co‐stimulatory molecules.
Nuclear factor‐κB and caspases co‐operatively regulate the activation and apoptosis of human macrophages
Authors: Hida A. , Kawakami A. , Nakashima T. , Yamasaki S. , Sakai H. , Urayama S. , Ida H. , Nakamura H. , Migita K. , Kawabe Y. , Eguchi K. 
Published: March 2003

SummaryAccumulating evidence suggests that macrophages function as major effector cells in the pathological process of various human diseases. We examined here the role of nuclear factor‐κB (NF‐κB) and caspases in the regulation of activation and apoptosis of macrophages. Activation of the human monoblastic leukaemia cell line, U937, by phorbol 12‐myristate 13‐acetate (PMA) increased the expression of CD14/CD86, and cytokine production. PMA stimulation also increased the expression of both pro‐caspase‐8 and pro‐caspase‐3 in U937, but not apoptosis or intracellular caspase‐3 activity. PMA also increased the expression of X‐chromosome‐linked inhibitor of apoptosis protein (XIAP) in U937, suggesting an inhibitory action for XIAP on the caspase cascade in PMA‐stimulated U937. Electrophoretic mobility shift assay (EMSA) showed a significant increase of nuclear NF‐κB activity in PMA‐stimulated U937. When a potent NF‐κB inhibitor, pyrrolidine dithiocarbamate (PDTC), was added to U937 cell culture in the presence of PMA, apoptosis was triggered by activation of caspase‐3, which was induced by caspase‐8 activation. XIAP expression was markedly suppressed in PMA‐treated U937 in the presence of PDTC. The inhibitors of caspase‐8 and caspase‐3 mostly inhibited apoptosis of U937 treated with PMA in the presence of PDTC. Furthermore, a phenotype of U937 treated with PMA and PDTC in the presence of caspase inhibitor was almost identical to that of unstimulated U937. Our results suggest that the signalling pathways involved in the activation and apoptosis of human macrophages could be co‐operatively regulated by the use of NF‐κB and caspase inhibitors, thus enabling the control of macrophage function and number.
Co‐option of endocytic functions of cellular caveolae by pathogens
Authors: Shin J.‐S. , Abraham S. N. 
Published: March 2003

SummaryIt is increasingly becoming clear that various immune cells are infected by the very pathogens that they are supposed to attack. Although many mechanisms for microbial entry exist, it appears that a common route of entry shared by certain bacteria, viruses and parasites involves cellular lipid‐rich microdomains sometimes called caveolae. These cellular entities, which are characterized by their preferential accumulation of glycosylphosphatidylinositol (GPI)‐anchored molecules, cholesterol and various glycolipids, and a distinct protein (caveolin), are present in many effector cells of the immune system including neutrophils, macrophages, mast cells and dendritic cells. These structures have an innate capacity to endocytoze various ligands and traffic them to different intracellular sites and sometimes, back to the extracellular cell surface. Because caveolae do not typically fuse with lysosomes, the ligands borne by caveolar vesicles are essentially intact, which is in marked contrast to ligands endocytozed via the classical endosome–lysosome pathway. A number of microbes or their exotoxins co‐opt the unique features of caveolae to enter and traffic, without any apparent loss of viability and function, to different sites within immune and other host cells. In spite of their wide disparity in size and other structural attributes, we predict that a common feature among caveolae‐utilizing pathogens and toxins is that their cognate receptor(s) are localized within plasmalemmal caveolae of the host cell.
Isolation, cloning and functional characterization of porcine mannose‐binding lectin
Authors: Agah A. , C M. , Montalto  , Young K. , Stahl G. L. 
Published: March 2003

SummaryBinding of mannose‐binding lectin (MBL), a C‐type lectin, and its associated serine proteases, MASP‐1 and MASP‐2, to cell surface carbohydrates activates the lectin complement pathway. As MBL plays an important role in innate immunity, it has been cloned and characterized in several species. While the pig may be used as a source of organs/tissues for xenotransplantation, little is known about its MBL, thus, we report the isolation of three monomeric forms of MBL from porcine serum. Sodium dodecyl sulphate–polyacrylamide gel electrophoresis and Coomassie staining of reduced porcine MBL revealed the presence of three monomeric forms with approximate molecular masses of 30 000, 32 000 and 34 000. Protein sequencing identified these monomeric forms as one single protein, suggesting post‐translational modification. Western blot analysis demonstrated the cross‐reactivity of anti‐human MBL polyclonal antibody with porcine MBL. A full‐length porcine liver MBL cDNA was isolated and the predicted amino acid sequence exhibited 64·9% identity with human MBL and 50·2% and 56·7% identity with rat A and C MBL, respectively. Furthermore, Northern blot analysis demonstrated the presence of a single (∼1·4–1·6 kilobase pair) transcript in porcine liver. Addition of purified porcine MBL to MBL‐deficient human sera augmented N‐acetylglucosamine inhibitable C3 deposition to mannan‐coated plates in a dose‐dependent manner. Taken together, these data demonstrate that porcine and human MBL are highly conserved, sharing structural and functional characteristics.
Long‐term in vivo depletion of functional CD4+ T lymphocytes from calves requires both thymectomy and anti‐CD4 monoclonal antibody treatment
Authors: Valdez R. A. , McGuire T. C. , Brown W. C. , Davis W. C. , Knowles D. P. 
Published: March 2003

Summary In vivo depletion of lymphocyte subsets is a direct approach used for dissection of the mechanisms of protective immunity. Long‐term in vivo depletion of bovine T lymphocyte subpopulations with monoclonal antibody (mAb) treatment alone has been difficult to achieve. The objective of this study was to determine whether both thymectomy and anti‐CD4 mAb treatment would optimize long‐term in vivo depletion of functional bovine CD4+ T lymphocytes. Calves were thymectomized and treated with high doses of anti‐CD4 mAb (approximately 5 mg/kg) over 4 days followed by subsequent lower doses (approximately 0·3 mg/kg) administered twice weekly for an additional 7 weeks. Depletion of CD4+ T lymphocytes from blood, spleen and peripheral lymph nodes was significantly improved in thymectomized calves compared to thymus‐intact anti‐CD4 mAb‐treated calves. Significant differences in percentages of CD4+ T lymphocytes between thymectomized and thymus‐intact calves were sustained for the duration of the 8‐week study. Depletion of CD4+ T lymphocytes from thymectomized calves resulted in complete abrogation of lymphoproliferative responses to ovalbumin. In addition, thymectomized calves treated with anti‐CD4 mAb had significantly reduced immunoglobulin G1 and no detectable immunoglobulin G2 ovalbumin‐specific antibody responses compared to thymus‐intact anti‐CD4 mAb‐treated calves. The results of this study demonstrate that both thymectomy and treatment with anti‐CD4 mAb are required for long‐term in vivo depletion of functional bovine CD4+ T lymphocytes.
Cleavage of E2F‐1‐regulating proteins and activation of E2F‐1 during CD95‐induced death of thymocytes
Authors: Bi BaoYuan , Littlewood Nicola K. , Crispe I. Nicholas 
Published: March 2003

SummaryThe CD95 death receptor activates caspases that cleave a variety of intracellular substrates, including cell cycle control proteins. However, the significance of this cleavage for the induction of apoptosis is unclear. In this study, CD95‐induced cleavage of the G1/S checkpoint regulator proteins, retinoblastoma protein (pRb) and murine‐double‐minute‐2 (mdm‐2), was associated with an increased protein concentration of a key transcription factor, E2F‐1, which is regulated by both of them. Furthermore, DNA‐binding activity to E2F sites is increased. In thymocytes, CD95‐induced apoptosis was associated with increased E2F‐1 DNA‐binding activity, while thymocytes that lacked E2F‐1 were less susceptible to CD95‐induced apoptosis. We conclude that the G1/S checkpoint is an important target of CD95 signalling. CD95‐activated caspases cleave regulator proteins to increase E2F‐1 activity, and inappropriate activation of E2F‐1 is part of the mechanism of CD95‐induced apoptosis.
A carbohydrate neoepitope that is up‐regulated on human mononuclear leucocytes by neuraminidase treatment or by cellular activation
Authors: Quinn Mark T. , Swain Steve D. , Parkos Charles A. , Jutila Kathryn L. , Siemsen Daniel W. , Kurk Sandra L. , Jesaitis Algirdas J. , Jutila Mark A. 
Published: March 2003

SummaryThe expression of cell‐surface antigens can delineate specific leucocyte developmental or functional stages. For example, certain membrane glycoproteins are expressed selectively on leucocyte subsets only after activation. Leucocyte activation can also induce changes in carbohydrate epitopes expressed on surface antigens. In the present studies, we report on a novel monoclonal immunoglobulin M antibody (mAb 13.22) that recognizes a unique carbohydrate epitope expressed on human leucocyte membrane proteins. Characterization of mAb 13.22 specificity by immunoblotting showed that it recognized proteins of MW ∼95 000 and 150 000, including both CD18 and CD11b. The mAb 13.22 epitope was removed by N‐glycosidase F but not by endoglycosidase H or fucosidase, demonstrating that it is an N‐linked carbohydrate antigen. Interestingly, immunoblot staining was enhanced after neuraminidase treatment, suggesting that the antibody epitope might also be partially masked by sialic acid. In resting leucocytes, the mAb 13.22 antigen was expressed strongly on neutrophils, while dull staining was present on monocytes, and no lymphocyte staining was observed. In marked contrast, treatment of leucocytes with neuraminidase resulted in exposure of a mAb 13.22 neoepitope on a subset of lymphocytes (primarily T lymphocytes and natural killer cells) as well as up‐regulated staining more than 18‐fold on monocytes. Activation of lymphocytes in culture with phytohaemagglutinin or concanavalin A also unmasked the mAb 13.22 neoepitope on ∼37% of the CD45RO+ lymphocytes. Furthermore, analysis of leucocytes collected from the synovial fluid of patients with rheumatoid arthritis showed that ∼18% of the lymphocytes present expressed the mAb 13.22 neoepitope. Taken together, our results suggest that the mAb 13.22 carbohydrate neoepitope could represent a physiologically relevant marker that is up‐regulated on leucocyte subsets during the inflammatory response.