bluuebunny/crossref_metadata_2025 huggingface dataset:
The intercellular adhesion molecule type‐1 is required for rapid activation of T helper type 1 lymphocytes that control early acute phase of genital chlamydial infection in mice
Authors: Igietseme J. U. , Ananaba G. A. , Bolier J. , Bowers S. , Moore T. , Belay T. , Lyn D. , Black C. M.
Published: March 2003

Recent studies in animal models of genital chlamydial disease revealed that early recruitment of dendritic cells and specific T helper type‐1 (Th1) cells into the genital mucosae is crucial for reducing the severity of the acute phase of a cervico‐vaginal infection and arresting ascending disease. These immune effectors are therefore important for preventing major complications of genital chlamydial infection. Other in vitro studies showed that intercellular adhesion molecule‐1 (ICAM‐1) plays a role in the antichlamydial action of specific CD4+ and CD8+ T cells. In the present study, we investigated the clinicopathological consequences of ICAM‐1 deficiency during chlamydial genital infection in ICAM‐1 knockout (ICAM‐1KO) mice, and analysed the cellular and molecular immunological bases for any observed pathology or complication. Following a primary genital infection of female ICAM‐l–/– and ICAM‐1+/+ mice, the intensity of the disease during the first 3 weeks (as assessed by shedding of chlamydiae in the genital tract) was significantly greater in ICAM‐1KO mice than in ICAM‐1+/+ mice (P < 0·0001), although both ICAM‐l–/– and ICAM‐1+/+ mice subsequently cleared the primary infection. There was greater ascending disease during the initial stage of the infection, and a higher incidence of tubal disease (hydrosalpinx formation) after multiple infections in ICAM‐l–/– mice. Analysis of the cellular and molecular bases for the increased acute and ascending disease in ICAM‐l–/– mice revealed that the high affinity of ICAM‐1 for leucocyte function antigen type‐1 is a property that promotes rapid activation of specific Th1 cells, as well as their early recruitment into the genital mucosa. Moreover, ICAM‐1 was more important for naive T‐cell activation than primed Th1 cells, although its absence delayed or suppressed immune T‐cell activation by at least 50%. Taken together, these results indicated that ICAM‐1 is crucial for rapid T‐cell activation, early recruitment and control of genitally acquired Chlamydia trachomatis.
Inflammatory cytokine production by immunological and foreign body multinucleated giant cells
Authors: Hernandez‐Pando R. , Bornstein Q. L. , Aguilar Leon D. , Orozco E. H. , Madrigal V. K. , Martinez Cordero E.
Published: March 2003

Multinucleated giant cells (MGC) are a common feature of granulomas. The mechanism of their formation has been studied extensively, but their function has not been completely characterized. A new method for the in vivo production of MGC was developed involving subcutaneous injection of microscopic nitrocellulose particles with adsorbed mycobacterial antigens into the footpads of sensitized BALB/c mice (immune [I]‐MGC), or by nitrocellulose administration to non‐sensitized mice (foreign body [FB]‐MGC). The development of granulomas with a highly enriched MGC population was observed 2 weeks after the nitrocellulose injection. MGC were larger with a greater number of nuclei in I‐MGC than in FB‐MGC. From days 7–28 after nitrocellulose administration, the production of interleukin‐1α (IL‐1α) and tumour necrosis factor‐α (TNF‐α) was demonstrated in both MGC types by in situ reverse transcription–polymerase chain reaction (RT–PCR) and immunohistochemistry. After 2 months, the MGC had ceased production of IL‐1α and TNF‐α, but the expression of transforming growth factor‐β (TGF‐β) was very high, occurring together with extensive fibrosis. These results suggest that MGC are an active source of inflammatory cytokines, which can contribute to the initiation, maintenance and down‐regulation of granulomatous inflammation induced by immunological and inert substances.
Comparative analysis of integrin expression on monocyte‐derived macrophages and monocyte‐derived dendritic cells
Authors: Ammon C. , Meyer S. P. , Schwarzfischer L. , Krause S. W. , Andreesen R. , Kreutz M.
Published: March 2003

Both macrophages (MAC) and dendritic cells (DC) are members of the mononuclear phagocyte system (MPS) with monocytes (MO) as common precursor cells. Cells of the MPS are able to take up, process and present antigens to T lymphocytes, thereby inducing a primary or secondary immune response. Adhesion molecules are of crucial importance for the interaction of antigen‐presenting cells with immune cells, especially T lymphocytes. By representational difference analysis, we identified CD49c (VLA‐3), a member of the β1‐integrin family of adhesion receptors, as differentiation‐associated antigen in MO‐derived MAC. In contrast, MO‐derived DC did not express CD49c mRNA. These data prompted us to compare the integrin expression pattern of MAC and DC. Both cell types showed a low expression of the α‐chains of the β1‐integrins CD49a, CD49b, CD49d and CD49e, whereas a marked difference was observed for CD49c and CD49f. Expression of both integrins increased during MO to MAC differentiation, but was not detectable on DC. In parallel the β1‐chain (CD29) was clearly up‐regulated during MO to MAC differentiation but was only weakly expressed on DC. On the other hand, the β2‐integrins CD11a, CD11b, CD11c and CD18 were all expressed on MAC and DC. Beside their role in cell–cell interaction and adhesion, β2‐integrins are also known as possible binding molecules for bacteria and lipopolysaccharide (LPS), especially for high LPS concentrations. Therefore we investigated the LPS response of MAC versus DC in terms of tumour necrosis factor‐α (TNF‐α) release. DC were less responsive to low doses of LPS, which can easily be explained by the very low CD14 expression on DC compared for MAC. In contrast, the TNF‐α response was comparable to MAC when DC were stimulated with high LPS concentrations. Our results show a specific, differentiation‐dependent pattern of β1‐ and β2‐integrin expression on in vitro‐generated MAC and DC. We suggest that the high expression of CD11/CD18 on DC could be involved in the LPS binding of DC. As LPS is not only an activation but also a differentiation stimulus for DC, the expression of CD11/CD18 on DC may be important for the successful maturation of DC and thereby the initiation of a primary immune response.
Engineering the microflora to vaccinate the mucosa: serum immunoglobulin G responses and activated draining cervical lymph nodes following mucosal application of tetanus toxin fragment C‐expressing lactobacilli
Authors: Shaw D. M. , GaerthÉ B. , Leer R. J. , Van Der Stap J. G. M. M. , Smittenaar C. , Heijne Den Bak‐Glashouwer M.‐J. , Thole J. E. R. , Tielen F. J. , Pouwels P. H. , Havenith C. E. G.
Published: March 2003

The delivery of antigens to mucosal‐associated lymphoid tissues in paediatric and immunocompromised populations by safe, non‐invasive vectors, such as commensal lactobacilli, represents a crucial improvement to prevailing vaccination options. In this report, we describe the oral and nasal immunization of mice with vaccines constructed through an original system for heterologous gene expression in Lactobacillus in which the 50 000‐molecular weight (MW) fragment C of tetanus toxin (TTFC) is expressed either as an intracellular or a surface‐exposed protein. Our data indicate that L. plantarum is more effective in this respect than L. casei and that, under the experimental conditions investigated, delivery of TTFC expressed as an intracellular antigen is more effective than cell‐surface expression. Immunization of mice with live recombinant lactobacilli induced significant levels of circulating TTFC‐specific immunoglobulin G (IgG) following nasal or oral delivery of vaccine strains. In addition, following nasal delivery, secretory immunoglobulin A (sIgA) was induced in bronchoalveolar lavage fluids, as were antigen‐specific antibody‐secreting cells and antigen‐specific T‐cell activation in draining lymph nodes, substantiating their potential for safe mucosal delivery of paediatric vaccines.
Staphylococcal enterotoxin B induces potent cytotoxic activity by intraepithelial lymphocytes
Authors: Roberts A. I. , Blumberg R. S. , Christ A. D. , Brolin R. E. , Ebert E. C.
Published: March 2003

In food poisoning, Staphylococcus aureus secretes staphylococcal enterotoxin B (SEB), a superantigen that causes intense T‐cell proliferation and cytotoxicity. The effects of SEB on lytic activity by human intestinal intraepithelial lymphocytes (IEL) were investigated. Jejunal IEL, from morbidly obese individuals undergoing gastric bypass operations, were tested for SEB‐induced cytotoxicity against C1R B‐lymphoblastoid cells, HT‐29 adenocarcinoma cells, or CD1d‐transfected cells using the 51Cr‐release assay. Fas and Fas ligand expression were detected by immunofluorescence and flow cytometry and soluble ligand by enzyme‐linked immunosorbent assay (ELISA). In the presence of SEB, IEL became potently cytotoxic against C1R cells and interferon‐γ (IFN‐γ)‐precultured HT‐29 cells, causing 55 ± 10% and 31 ± 6% lysis, respectively, greater than that by phytohaemagglutinin (PHA)‐, interleukin‐2 (IL‐2)‐, or anti‐T‐cell receptor (TCR)‐activated IEL. SEB‐stimulated peripheral blood (PB) CD8+ T cells lysed similar numbers of C1R cells but fewer HT‐29 cells (53 ± 13% and 8 ± 5%, respectively). IEL killing of C1R cells involved interaction of major histocompatibility complex (MHC) class II with TCR, CD2 with CD58, and CD11a with CD54, and was perforin mediated. SEB‐induced IEL lysis of HT‐29 cells, in contrast, was caused by an unknown target cell structure, not MHC class II or CD1d, and resulted from a combination of perforin and Fas‐mediated events. The potent cytotoxic activities of IEL promoted by SEB utilize two different mechanisms, depending on the surface receptors expressed by the target cells.
Professional and non‐professional antigen‐presenting cells in the porcine small intestine
Authors: Haverson K. , Singha S. , Stokes C. R. , Bailey M.
Published: March 2003

We have previously presented evidence of a highly organized and compartmentalized structure of the small intestinal lamina propria of the pig. In this work, we have identified at least two major populations of cells in this site expressing high levels of major histocompatibility complex (MHC) class II antigens. One is CD45 positive and is a potent initiator of a primary immune response, this is a function usually associated with dendritic cells. These cells have characteristic dendritic morphology, but show evidence of phagocytosis as well as other phenotypic markers of immature dendritic cells. Some cells show evidence of ongoing immune maturation. We have also isolated CD45 negative endothelial cells bearing significant amounts of MHC class II, which do not trigger a mixed lymphocyte reaction. These findings have implications for the functional role of healthy gut lamina propria and clearly implicate this site as capable of differential antigen presentation by a heterogeneous population of antigen‐presenting cells.
Cellular but not humoral immune responses generated by vaccination with dendritic cells protect mice against leukaemia
Authors: Colombo B. M. , Lacave R. , Pioche‐durieu C. , Masurier C. , Lemoine F. M. , Guigon M. , Klatzmann D.
Published: March 2003

Dendritic cells (DC) are extremely efficient at generating both prophylactic and therapeutic anti‐tumour immunity. We aimed to analyse the respective roles of humoral and cellular immune responses generated in mice vaccinated with bone marrow (BM)‐derived DC in terms of in vivo anti‐leukaemia effect. We used the murine L1210 B lymphocytic leukaemia genetically modified to express on the cell surface of human CD4 (hCD4) (L1210/hCD4) as a model tumour‐associated antigen (TAA). DC cultures were loaded with either purified soluble hCD4 (shCD4) protein or unfractionated L1210/hCD4 extracts and injected as vaccine into mice. The efficacy of these vaccinations was compared with that of vaccination with shCD4 protein emulsified in Freund’s adjuvant (FA). We evaluated the immune responses generated after these vaccinal protocols and the survival rate of vaccinated mice subsequently challenged with a lethal injection of L1210/hCD4 cells. Our results demonstrated that vaccination with shCD4 protein or tumour extract‐loaded DC mainly generated an hCD4 antigen‐specific cell‐mediated cytotoxic immune response that was associated with a specific protection against leukaemia. In contrast, vaccination with the protein emulsified in FA only generated potent humoral immune responses that were not protective against leukaemia. Altogether, our results indicate that the unique property of loaded DC to trigger an anti‐leukaemia protective effect is mainly associated with cellular immune responses.
Interferon‐γ plays a critical role in intestinal immunity against Salmonella typhimurium infection
Authors: Bao S. , Beagley K. W. , France M. P. , Shen J. , Husband A. J.
Published: March 2003

Salmonella bacteria are a major cause of food‐borne infectious diarrhoea and there is great interest in understanding the pathogenesis of Salmonella infection and in vaccine development. Potential vaccines include the aromatic mutants of S. typhimurium. Such non‐lethal Aro mutants have also been useful for studying Salmonella infections in mouse models. Studies of systemic infection, using these Aro mutants, in both normal and cytokine gene knockout mice, indicate that interferon‐γ (IFN‐γ) plays a key role in the resolution of Salmonella infection. The present studies have investigated the outcome of oral infection in mice with attenuated Salmonella because this infection route mimics natural infection in humans. In IFN‐γ gene knockout (IFN‐γ–/–) mice, intestinal immunity was impaired and oral challenge resulted in disseminated septicaemia 2 weeks later. No dissemination of infection was seen in wild‐type mice. In wild‐type mice, both CD4 and CD8 cell numbers increased in the gut following Salmonella challenge, together with increased expression of major histocompatibility complex (MHC) II and vascular cell adhesion molecule‐1 (VCAM‐1). No such changes were seen in IFNγ–/– mice. Following oral challenge, antilipopolysaccharide (LPS) and antiphosphoryl choline antibodies increased by more than 100‐fold in both serum and faecal pellet extracts of IFNγ–/– mice compared with wild‐type mice. Our data show that IFN‐γ production is essential for resolution of enteric Salmonella infection and that antibody has little effect on this process.
Redundancy or cell‐type‐specific regulation? Tumour necrosis factor in alveolar macrophages and mast cells
Authors: Déry R. E. , Lin T.‐J. , Befus A. D. , Milne C. D. , Moqbel R. , Ménard G. , Bissonnette E. Y.
Published: March 2003

Tumour necrosis factor (TNF) is an important inflammatory cytokine produced by several cell types. To test the hypothesis that there is cell‐type‐specific regulation and not redundancy of TNF production, we investigated its production by alveolar macrophages (AM) and peritoneal mast cells (PMC). Cell lysates of freshly isolated AM and PMC contained 9 ± 3 pg and 57 ± 17 pg of TNF/106 cells, respectively. Furthermore, unstimulated PMC expressed 4 × 103‐fold more attomols of TNF mRNA/µg total RNA compared with AM. These data may explain in part the greater TNF‐dependent cytotoxicity of PMC. Furthermore, fixed PMC showed significantly higher TNF‐dependent cytotoxic activity than AM (sevenfold), suggesting that PMC express more membrane TNF than AM. Although AM and PMC contain different amounts of TNF, antigen stimulation caused a similar release of TNF from sensitized rats. Interferon (IFN)‐γ, respectively, stimulated and inhibited AM and PMC TNF‐dependent cytotoxicity whereas lipopolysaccharide (LPS) significantly stimulated TNF‐dependent cytotoxicity in both cell types. However, TNF released (AM 400‐fold and PMC threefold) and TNF mRNA expression, as measured by competitive reverse transcription–polymerase chain reaction (AM 7 × 103‐fold and PMC twofold), were considerably greater in LPS‐stimulated AM than PMC. Our data indicate that TNF is differentially expressed in these two cell types and that its production is dependent on the nature of the stimulus. These data provide vital basis in experimental approaches aimed at modulating the effect of TNF in airway disease conditions involving both AM and mast cells.
Intercellular communication in the immune system: differential expression of connexin40 and 43, and perturbation of gap junction channel functions in peripheral blood and tonsil human lymphocyte subpopulations
Authors: Oviedo‐orta E. , Hoy T. , Evans W. H.
Published: March 2003

The distribution and function of connexins (integral membrane proteins assembled into gap junction intercellular communication channels) were studied in human lymphocyte subpopulations. The expression of mRNA encoding connexins in peripheral blood and tonsil‐derived T, B and natural killer (NK) lymphocytes was examined. Connexin43 (Cx43) mRNA was expressed in peripheral blood and tonsil lymphocytes, but Cx40 mRNA expression was confined to tonsil‐derived T and B lymphocytes; Cx26, Cx32, Cx37 and Cx45 were not detected by reverse transcription–polymerase chain reaction (RT–PCR). Western blot analysis also demonstrated the presence of Cx40 and Cx43 proteins in T and B lymphocytes in a manner coincidental to the mRNA detection. Stimulation in vitro of T and B lymphocytes with phytohaemagglutinin (PHA) and lipopolysaccharide (LPS), respectively, increased Cx40 and Cx43 protein expression. Flow cytometric analysis, using antibodies to extracellular loop amino acid sequences of connexins, confirmed the surface expression of connexins in all lymphocyte subpopulations. Assembly of connexins into gap junctions providing direct intercellular channels linking attached lymphocytes was demonstrated by using a dye transfer technique. The exchange of dye between lymphocytes was inhibited by a connexin extracellular loop mimetic peptide and α‐glycyrrhetinic acid, two reagents that restrict intercellular communication across gap junctions. Dye coupling occurred between homologous and heterologous co‐cultures of T and B lymphocytes, and was not influenced by their stimulation with PHA and LPS. The connexin mimetic peptide caused a significant decrease in the in vitro synthesis of immunoglobulin M (IgM) by T‐ and B‐lymphocyte co‐cultured populations in the presence or absence of stimulation by PHA. The results identify connexins as important cell surface components that modulate immune processes.
Functional dissection of the cytoplasmic subregions of the interleukin‐5 receptor α chain in growth and immunoglobulin G1 switch recombination of B cells
Authors: Moon B.‐G. , Yoshida T. , Shiiba M. , Nakao K. , Katsuki M. , Takaki S. , Takatsu K.
Published: March 2003

The interleukin‐5 receptor α chain (IL‐5Rα) is known to regulate the development and function of B cells and eosinophils. Although the functions of IL‐5Rα cytoplasmic domain subregions have been studied extensively using cultured cell lines, this approach has limitations when studying the functions of distinct primary B‐cell subpopulations and their responsiveness to IL‐5. In the present study, we generated mice on an IL‐5Rα null background, each expressing a mutant form of an IL‐5Rα transgene ligated to a µ enhancer and VH promoter, either lacking the cytoplasmic DC3 region or substituting two proline residues for alanine (ApvA) in the membrane‐proximal ppvp motif of the cytoplasmic domain. The ppvp motif, which mediates activation of JAK2/STAT5 and Btk, also contributes to c‐fos, c‐jun and c‐myc expression. IL‐5Rα null mutant mice showed impaired B‐1‐cell development, reduced serum immunoglobulin G3 (IgG3) and IgM, no IL‐5‐induced enhancement of B‐cell proliferation and IL‐5‐induced switch recombination from the µ gene to γ1 gene; these were not recovered following the expression of the ApvA mutant. In contrast, absence of the DC3 region affected the IL‐5‐induced switch recombination from the µ to the γ1 gene and B‐1‐cell development, while IL‐5‐induced proliferation and IgM production were at levels similar to those of B cells expressing wild‐type IL‐5Rα transgene. The results clearly indicated that the ppvp motif and the DC3 region of IL‐5Rα played distinct roles in B‐cell proliferation and differentiation. Thus, this present approach offers new insights into the functions of the cytoplasmic subregions of IL‐5Rα, in particular its carboxy‐terminal region.
Role of oestrogen receptors α and β in immune organ development and in oestrogen‐mediated effects on thymus
Authors: Erlandsson Malin C. , Ohlsson Claes , Gustafsson Jan‐Åke , Carlsten Hans
Published: March 2003

Oestrogens affect the development and regulation of the immune system. To determine the role of oestrogen receptors α (ER‐α) and β (ER‐β) on the development of the immune system, male ER‐α (ERKO) and ER‐β (BERKO) mice, as well as αβ‐double knockout (DERKO) mice, were studied. Deletion of ER‐α led to hypoplasia of both thymus and spleen. Interestingly, a higher frequency of immature double CD4+ CD8+ thymocytes was found in ER‐α− mice compared with ER‐α+ mice. Female oophorectomized BERKO mice given oestradiol (E2) displayed a similar degree of thymic atrophy compared with the wild‐type strain but showed only limited involution of thymus cortex and no alteration of thymic CD4/CD8 phenotype expression. Our data demonstrate that expression of ER‐α, but not ER‐β, is mandatory in males for development of full‐size thymus and spleen, whereas expression of ER‐β is required for E2‐mediated thymic cortex atrophy and thymocyte phenotype shift in females. A potential background for the above findings may be down‐regulated activity in the growth hormone/insulin‐like growth factor‐1 (GH/IGF‐1) axis in males lacking ER‐α and suppressed sensitivity of females lacking ER‐β to E2‐mediated suppression of IGF‐1.
Acquisition of immune function during the development of the Langerhans cell network in neonatal mice
Authors: Dewar Andrea L. , Doherty Kathleen V. , Woods Gregory M. , Lyons A. Bruce , Muller H. Konrad
Published: March 2003

The immunological function of the Langerhans cell (LC) network in neonatal skin was examined by defining the development of cutaneous immunity relative to the structure, phenotype and function of the epidermal LC network in neonatal, juvenile and adult mice. Analysis of epidermal sheets showed the presence of major histocompatibility complex (MHC) II+, multilectin receptor DEC‐205– cells within the epidermis of 3‐day‐old mice; both cell density and DEC‐205 expression increased until day 14. When visualized with antibodies directed at MHC II, the network was poorly formed in 3‐ and 7‐day‐old mice, as there was a lower cell density and poor MHC II expression on dendritic processes, compared to mice at day14. Application of a fluorescent antigen to 3‐day‐old mice revealed that the LC were inefficient in transporting antigen to the draining lymph node. There was an improvement at day 7 and by day 14 comparable numbers of antigen carrying cells were detected in the lymph nodes of 6‐week‐old mice. The reduced antigen carriage in 3‐ and 7‐day‐old mice correlated with a poor contact sensitivity response. This was not simply due to failure to present antigen, but development of immunosuppression, as transfer of T cells from adult mice that were previously treated with antigen when they were 3 days old, to adult recipients resulted in antigen specific immunosuppression. Analysis of CD80 and CD86 expression showed that LC from day 3 skin expressed CD80, but not CD86 and application of antigen through this skin was inefficient in upregulating CD86. These findings indicate that when the neonatal LC network is poorly developed it is functionally immature and antigen applied through this ‘functionally immature network’ results in antigen specific immunosuppression.
Maternal tolerance is not critically dependent on interleukin‐4
Authors: Bonney Elizabeth A.
Published: March 2003

Pregnant animals can generate and maintain immune responses to fetal antigens. This however, does not usually lead to fetal loss. At least two types of immune response are recognized. T helper type 1 (Th1) responses support the generation of cellular cytotoxicity. In contrast, Th2‐type responses support the production of non‐cytotoxic antibody and suppress the Th1‐type. One attempt to explain why the fetus is not generally rejected has been to suggest that during pregnancy Th2‐type responses are dominant. These responses rely heavily on interleukin‐4 (IL‐4) for both functions. This work focuses on maternal immunity to the male antigen H‐Y, which is expressed in male fetuses. When injected with male spleen cells, female mice of certain strains mount a cytotoxic immune response to H‐Y. However, pregnant females immunized in this way do not deliver litters with fewer males. To help delineate the possible role of IL‐4 in such maternal tolerance, female mice genetically deficient in IL‐4 were studied. The results show that: (1) deficiency in maternal IL‐4 does not affect fertility, (2) deficiency in IL‐4 is not associated with selective loss of male offspring in unimmunized mice, (3) pregnancy does not obliterate anti‐H‐Y reactivity in immunized mice and (4) maternal immunity to H‐Y in the absence of IL‐4 does not result in loss of male offspring. The results suggest that IL‐4‐dependent Th2‐type responses are not critical to maternal tolerance. Other cytokines must be examined for their role in this phenomenon.
Nitric oxide protects cultured rheumatoid synovial cells from Fas‐induced apoptosis by inhibiting caspase‐3
Authors: Migita Kiyoshi , Yamasaki Satoshi , Kita Masako , Ida Hiroaki , Shibatomi Kazutaka , Kawakami Atsushi , Aoyagi Takahiko , Eguchi Katsuni
Published: March 2003

Nitric oxide (NO) is elevated in the synovial fluids and sera of patients with rheumatoid arthritis (RA) and is thought to be an important proinflammatory mediator in the rheumatoid synovium. To test the hypothesis that NO might modulate the apoptosis‐inducing signal pathway, we investigated the effects of NO on rheumatoid synovial‐cell apoptosis induced by Fas ligation with anti‐Fas antibody. Pretreatment of synovial cells with the NO donor S‐nitro‐N‐acetylpenicillamine (SNAP) prevented the Fas‐mediated induction of apoptosis. The activation of caspase‐3 was required to mediate Fas‐induced synovial cell apoptosis. The NO donor SNAP inhibited Fas‐induced caspase‐3 activation in rheumatoid synovial cells. However, NO did not interrupt Fas‐induced caspase‐8 cleavage or subsequent cytochrome c release into the cytosol in rheumatoid synovial cells. These data indicate that NO prevents apoptosis in rheumatoid synovial cells by directly inhibiting caspase‐3 activation. Thus, we propose that NO interferes with cell death signal transduction and may contribute to rheumatoid synovial cell proliferation by inhibiting induction of apoptosis.
Leucocyte recruitment during enteric nematode infection
Authors: McDermott Jacqueline R. , Grencis Richard K. , Else Kathryn J.
Published: March 2003

Resolution of infection with the intestinal nematode Trichinella spiralis depends on the host mounting a T helper 2 (Th2) response. It is known that both mast cells and T cells play a crucial role. We have previously shown that efficient migration of mast cells to the gut during infection depends on their expression of the integrin β7. β7 forms a heterodimer complex with either αE or α4 integrin chains, αEβ7 binding to E‐cadherin expressed by epithelial cells and α4β7 binding to mucosal addressin cell adhesion molecule (MAdCAM‐1) on the endothelium. We were interested to know whether dysfunctional mast cell localization to the gut in the absence of β7 was due to the failure of α4β7 to bind to MAdCAM‐1 or the failure of αEβ7 to bind to E‐cadherin. We used blocking monoclonal antibodies against αE (M290) or α4 (PS2) or β7 (HB293) during T. spiralis infection of C57BL/6 mice and found that all antibody treatments reduced mastocytosis. In contrast, none of the antibody treatments prevented the migration of CD3+ T cells into the intestine. These results indicate that during inflammation (a) there is integrin redundancy for lymphocytes but not for mast cells and (b) both αEβ7 and α4β7 are crucial either for the entry of mast cells into the gut or for their maturation once they have entered.
Acholeplasma laidlawii up‐regulates granulysin gene expression via transcription factor activator protein‐1 in a human monocytic cell line, THP‐1
Authors: Kida Yutaka , Kuwano Koichi , Zhang Ye , Arai Sumio
Published: March 2003

An antimicrobial protein granulysin is constitutively expressed in cytotoxic T lymphocytes (CTL) and natural killer (NK) cells. However, little is known about the precise regulatory mechanisms underlying granulysin gene expression. In this study, we examined the regulatory mechanisms underlying granulysin gene expression using a human monocytic cell line, THP‐1, treated with Acholeplasma laidlawii. The level of granulysin mRNA expression in THP‐1 cells was significantly augmented in response to stimulation with A. laidlawii. The transfection of reporter gene constructs into THP‐1 cells indicated that DNA sequences between residues −329 and −239, relative to the transcriptional start site of the granulysin gene, are responsible for mediating gene induction. In addition, mutagenesis of a putative activator protein‐1 (AP‐1)‐binding site between residues −277 and −271 in the granulysin promoter resulted in the reduction of granulysin promoter activity. Electrophoretic mobility shift assays (EMSA) demonstrated that nuclear extract prepared from A. laidlawii‐treated THP‐1 cells can generate specific binding to DNA oligonucleotides encompassing the AP‐1‐binding site, whereas unstimulated nuclear extract from the cells failed to do so. Furthermore, competition and supershift assays confirmed that A. laidlawii can induce the activation of AP‐1. These results indicate that AP‐1 dominantly participates in the regulation of inducible granulysin gene expression in THP‐1 cells. Therefore, the finding of inducible granulysin gene expression by A. laidlawii suggests that inducible granulysin in macrophages may function as a protective weapon when microbial invasion occurs.
Chemokine signalling: pivoting around multiple phosphoinositide 3‐kinases
Authors: Curnock Adam P. , Logan Marisa K. , Ward Stephen G.
Published: March 2003

The role of chemokines in mediating directional cell migration is well established, but more recently it has become evident that chemokines are able to couple to distinct signalling pathways that are involved in not only chemotaxis, but also cell growth and transcriptional activation. The signalling pathway controlled by the phosphoinositide 3‐kinase (PI3K) family of lipid kinases has been the focus of much attention with respect to their role in chemokine‐mediated functional responses. Indeed, there now exists convincing biochemical, pharmacological and genetic evidence that both CC and CXC chemokines stimulate PI3K‐dependent chemotaxis of inflammatory cells such as eosinophils, macrophages, neutrophils and T lymphocytes. This review considers the role of individual PI3Ks (e.g. the p85/p110 heterodimer, PI3Kγ and PI3KC2α) as well their downstream effector targets in mediating chemokine‐stimulated cell migration.
Distribution of IFN‐γ, IL‐4 and TNF‐α protein and CD8 T cells producing IL‐12p40 mRNA in human lung tuberculous granulomas
Authors: Fenhalls Gael , Stevens Liesel , Bezuidenhout Juanita , Amphlett Gillian E. , Duncan Ken , Bardin Philip , Lukey Pauline T.
Published: March 2003

In order to examine the immune response at the site of pathology in tuberculosis, we analysed cytokines present in lung granulomas, their associations with each other and with caseous necrosis as well as the phenotype of the cellular infiltrate. Paraffin‐embedded tissue from the lungs of seven patients with pulmonary tuberculosis was analysed by immunohistochemistry and in situ hybridization to detect interferon‐γ (IFN‐γ), tumour necrosis factor‐α (TNF‐α) and interleukin‐4 (IL‐4) proteins and IL‐12p40 mRNA. All seven patients had granulomas staining positive for IFN‐γ, TNF‐α and IL‐12p40, but only four stained positive for IL‐4. Cells with the morphology of lymphocytes, macrophages and giant cells expressed TNF‐α, IFN‐γ and IL‐4 protein. Furthermore, CD68‐positive myeloid cells expressed IL‐12p40 mRNA, as expected, but a subset of CD3‐positive lymphocytes also expressed this mRNA. These lymphocytes producing IL‐12p40 also stained positive for CD8 but not CD4. A total of 141 granulomas were scored for the presence or absence of cytokine or necrosis and two major associations were identified. The first association was between IFN‐γ and IL‐12, with 76% of granulomas staining positive for both cytokines. Unexpectedly, those granulomas positive for IL‐4 were always positive for IFN‐γ. The second association was between TNF‐α and caseous necrosis, where all necrotic granulomas were TNF‐α positive. This association was modulated by IL‐4. Therefore, heterogeneity of cellular infiltrate and cytokine expression is observed between adjacent granulomas in the same patient.
A cathelicidin family of human antibacterial peptide LL‐37 induces mast cell chemotaxis
Authors: Niyonsaba François , Iwabuchi Kazuhisa , Someya Akimasa , Hirata Michimasa , Matsuda Hiroshi , Ogawa Hideoki , Nagaoka Isao
Published: March 2003

The mast cell is one of the major effector cells in inflammatory reactions and can be found in most tissues throughout the body. During inflammation, an increase in the number of mast cells in the local milieu occurs, and such accumulation requires directed migration of this cell population. As it has previously been reported that the human cathelicidin‐derived antibacterial peptide, LL‐37, stimulates the degranulation of mast cells, we hypothesized that LL‐37 could be a mast cell chemotaxin. The present study shows that LL‐37 is a potent chemotactic factor for mast cells. The chemotactic response was dose‐dependent and bell‐shaped, reaching an optimal concentration of 5 µg/ml. In addition, checkerboard analysis showed that cell migration towards this peptide was chemotactic rather than chemokinetic. Moreover, Scatchard analysis using 125I‐labelled LL‐37‐derived peptide revealed that LL‐37 has at least two classes of receptors, namely high‐ and low‐affinity receptors, on mast cells. Furthermore, the competitive binding assay suggested that LL‐37 is unlikely to utilize formyl peptide receptor‐like 1 (FPRL1), a functional LL‐37 receptor for neutrophil and monocyte migration, on mast cells. In addition, the treatment of cells with pertussis toxin and phospholipase C inhibitor, U‐73122, inhibited LL‐37‐mediated migration, indicating that LL‐37 induces mast cell chemotaxis through a Gi protein‐phospholipase C signalling pathway. These results show that besides its antibacterial activities, LL‐37 may have the potential to recruit mast cells to inflammation foci.
T‐cell dynamics after high‐dose chemotherapy in adults: elucidation of the elusive CD8+ subset reveals multiple homeostatic T‐cell compartments with distinct implications for immune competence
Authors: Fagnoni Francesco F. , Lozza Laura , Zibera Carlo , Zambelli Alberto , Ponchio Luisa , Gibelli Nadia , Oliviero Barbara , Pavesi Lorenzo , Gennari Roberto , Vescovini Rosanna , Sansoni Paolo , Da Prada Gianantonio , Robustelli Della Cuna Gioacchino
Published: March 2003

Recovery of total T cell numbers after in vivo T‐cell depletion in humans is accompanied by complex perturbation within the CD8+ subset. We aimed to elucidate the reconstitution of CD8+ T cells by separate analysis of putative naïve CD95− CD28+, memory CD95+ CD28+ and CD28− T cell compartments after acute maximal depletion by high‐dose chemotherapy (HD‐ChT) in women with high‐risk breast cancer. We found that recovery of putative naïve CD8+ CD95− CD28+ and CD4+ CD95− CD28+ T cells, was compatible with a thymus‐dependent regenerative pathway since their recovery was slow and time‐dependent, their values were tightly related to each other, and their reconstitution patterns were inversely related to age. By analysing non‐naïve T cells, a striking diversion between putative memory T cells and CD28− T cells was found. These latter increased early well beyond normal values, thus playing a pivotal role in total T‐cell homeostasis, and contributed to reduce the CD4 : CD8 ratio. In contrast, putative memory T cells returned to values not significantly different from those seen in patients at diagnosis, indicating that this compartment may recover after HD‐ChT. At 3–5 years after treatment, naïve T cells persisted at low levels, with expansion of CD28− T cells, suggesting that such alterations may extend further. These findings indicate that CD28− T cells were responsible for ‘blind’ T‐cell homeostasis, but support the notion that memory and naïve T cells are regulated separately. Given their distinct dynamics, quantitative evaluation of T‐cell pools in patients undergoing chemotherapy should take into account separate analysis of naïve, memory and CD28− T cells.
Involvement of ERK, p38 and NF‐κB signal transduction in regulation of TLR2, TLR4 and TLR9 gene expression induced by lipopolysaccharide in mouse dendritic cells
Authors: An Huazhang , Yu Yizhi , Zhang Minghui , Xu Hongmei , Qi Runzi , Yan Xiaoyi , Liu Shuxun , Wang Wenya , Guo Zhenghong , Guo Jun , Qin Zhihai , Cao Xuetao
Published: March 2003

Toll‐like receptors (TLR) are sentinel receptors capable of recognizing pathogen‐associated molecule patterns (PAMP) such as lipopolysaccharide (LPS) and CpG‐containing oligonucleotides (CpG ODN). TLR2 and TLR4 are major receptors for Gram‐positive and Gram‐negative bacterial cell wall components, respectively. TLR9 is necessary for CpG signalling. LPS or CpG ODN can activate immature dendritic cells (DC) and induce DC maturation characterized by production of cytokines, up‐regulation of co‐stimulatory molecules, and increased ability to activate T cells. However, little is known regarding the regulation of TLR gene expression in mouse DC. In this study, we investigated the regulation of TLR2, TLR4 and TLR9 gene expression by LPS in murine immature DC. TLR2, TLR4 and TLR9 mRNA were up‐regulated following LPS stimulation. The up‐regulation of TLR9 expression coincided with significantly increased production of tumour necrosis factor‐α induced by LPS plus CpG ODN. While inhibition of extracellular signal‐related kinase and NF‐κB activation suppressed the up‐regulation of the expression of TLR2, TLR4 and TLR9 mRNA, inhibition of p38 kinase prevented the up‐regulation of TLR2 and TLR4 mRNA expression but enhanced the up‐regulation of TLR9 expression. These results demonstrated that TLR2, TLR4 and TLR9 gene expression was differently regulated by LPS in mouse immature DC. Up‐regulation of TLR2, TLR4 and TLR9 expression by LPS might promote the overall responses of DC to bacteria and help to explain the synergy between LPS and other bacterial products in the induction of cytokine production.
The 52 000 MW Ro/SS‐A autoantigen in Sjögren's syndrome/systemic lupus erythematosus (Ro52) is an interferon‐γ inducible tripartite motif protein associated with membrane proximal structures
Authors: Rhodes Davd A. , Ihrke Gudrun , Reinicke Anna T. , Malcherek Georg , Towey Michael , Isenberg David A. , Trowsdale John
Published: March 2003

The 52 000 MW Ro/SS‐A (Ro52) protein is a major target of autoantibodies in autoimmune conditions such as systemic lupus erythematosus and Sjögren's syndrome. Recent genomic and bioinformatic studies have shown that Ro52 belongs to a large family of related RING/Bbox/coiled‐coil (RBCC) tripartite motif proteins sharing overall domain structure and 40–50% identity at the amino acid level. Ro52 also has a B30.2 domain at the C‐terminus. Using the human genome draft sequence, the genomic organization of the Ro52 gene on human chromosome 11p15.5 has been deduced and related to the protein domain structure. We show that the steady‐state levels of Ro52 mRNA are normally very low but are induced by cell activation with interferon‐γ. In transient transfection of HeLa cells, epitope‐tagged Ro52 protein was localized to unidentified membrane proximal rod‐like structures. Using in vitro coupled transcription/translation followed by immunoprecipitation, the autoimmune response to Ro52 protein was investigated and two distinct interactions were resolved. The Ro52 C‐terminal B30.2 domain interacts with human immunoglobulin independently of antibody specificities. Sera derived from patients with Sjögren's syndrome and systemic lupus erythematosus, in addition, contained specific autoantibodies directed towards the rest of the Ro52 molecule. The majority of these autoimmune sera also immunoprecipitated the Ro52‐related molecule RNF15. A possible role for Ro52 protein in alterations of plasma membranes during cellular activation or apoptosis is discussed.
Effects of human interleukin‐18 and interleukin‐12 treatment on human lymphocyte engraftment in NOD‐scid mouse
Authors: Senpuku Hidenobu , Asano Toshihiko , Matin Khairul , Abdus Salam M. , Tsuha YUZO , Horibata Shigeo , Shimazu Yoshihito , Soeno Yuichi , Aoba Takaaki , Sata Tetsutaro , Hanada Nobuhiro , Honda Mitsuo
Published: March 2003

NOD/LtSz‐prkdcscid/prkdcscid (non‐obese diabetic‐severe combine immunodeficiency; NOD‐scid) mice grafted with human peripheral blood lymphoid cells have been used as an in vivo humanized mouse model in various studies. However, cytotoxic human T cells are induced in this model during immune responses, which gives misleading results. To assist in grafting of human lymphocytes without the induction of cytotoxic human T cells, we investigated the effects of T helper type 1 (Th1) and Th2 cytokines on human lymphocyte grafting and migration, as well as the production of immunoglobulin deposited in glomeruli and human immunodeficiency virus‐1 (HIV‐1) infection using NOD‐scid mice. Administration of interleukin‐18 (IL‐18) and IL‐12 enhanced the grafting of human CD4+ and CD8+ T cells in the mice, whereas co‐administration prevented grafting due to interferon‐γ‐dependent apoptosis. Immunoglobulin A (IgA) deposits were observed in mice treated with IL‐18 alone, but not in those given phosphate‐buffered saline, IL‐12 alone, or IL‐18 + IL‐12. A high rate of HIV infection was also observed in the IL‐18‐treated group. Together, these results indicate that IL‐18 may be effective for the grafting and migration of CD4+ and CD8+ T cells, except for the induction of apoptosis and regulation of class‐switching IgA. IL‐18‐administered NOD‐scid mice provide a useful small humanized model for the study of HIV infection and IgA nephropathy.
Tryptophan deprivation sensitizes activated T cells to apoptosis prior to cell division
Authors: Lee Geon Kook , Park Hyeon Jin , Macleod Megan , Chandler Phillip , Munn David H. , Mellor Andrew L.
Published: March 2003

Cells expressing indoleamine 2,3‐dioxygenase (IDO), an enzyme which catabolizes tryptophan, prevent T‐cell proliferation in vitro, suppress maternal antifetal immunity during pregnancy and inhibit T‐cell‐mediated responses to tumour‐associated antigens. To examine the mechanistic basis of these phenomena we activated naïve murine T cells in chemically defined tryptophan‐free media. Under these conditions T cells expressed CD25 and CD69 and progressed through the first 12 hr of G0/G1 phase but did not express CD71, cyclin D3, cdk4, begin DNA synthesis, or differentiate into cytotoxic effector cells. In addition, activated T cells with their growth arrested by tryptophan deprivation exhibited enhanced tendencies to die via apoptosis when exposed to anti‐Fas antibodies. Apoptosis was inhibited by caspase inhibitor and was not observed when T cells originated from Fas‐deficient mice. These findings suggest that T cells activated in the absence of free tryptophan entered the cell cycle but cell cycle progression ceased in mid‐G1 phase and T cells became susceptible to death via apoptosis, in part though Fas‐mediated signalling. Thus, mature antigen‐presenting cells expressing IDO and Fas‐ligand may induce antigen‐specific T‐cell tolerance by blocking T‐cell cycle progression and by rapid induction of T‐cell activation induced cell death in local tissue microenvironments.
Regulation of epithelial cell cytokine responses by the α3β1 integrin
Authors: Lubin Farah D. , Segal Miriam , Mcgee Dennis W.
Published: March 2003

Epithelial cells (EC) from various tissues can produce important cytokines and chemokines when stimulated by proinflammatory cytokines. These EC also receive signals from cell surface integrins, like the α3β1 integrin, which is important in cell migration and wound healing of epithelial monolayers. However, little is known of the effect of integrin signals on cytokine responses by EC. Colonic Caco‐2 cells treated with an anti‐α3 integrin antibody prior to stimulation with the proinflammatory cytokine interleukin (IL)‐1 yielded suppressed levels of mRNA and secreted IL‐6, IL‐8 and monocyte chemoattractant protein‐1 as compared to cells treated with normal mouse immunoglobulin G. Lung A549 cells also showed a similar suppression of cytokine secretion. Likewise, treatment of the Caco‐2 cells with the same antibody suppressed tumour necrosis factor‐α‐stimulated IL‐6 secretion. Fab fragments of the anti‐α3 integrin antibody did not induce the suppressive effect but did block the suppressive effect of the whole antibody suggesting that the effect of the antibody required cross‐linking of the integrins. Finally, culture of the Caco‐2 cells on laminin type 5 (the major ligand for this integrin) yielded depressed levels of IL‐1‐induced IL‐6 secretion as compared to cells on laminin type 1. These data are the first indication that the α3β1 integrin may cause a suppression of cytokine responses by EC which may be important in regulating the capacity of EC to respond during inflammation or wound healing.
Neither lymphotoxin α nor lymphotoxin β receptor expression is required for biogenesis of lymphoid aggregates or differentiation of natural killer cells in the pregnant mouse uterus
Authors: Kather Angela , Chantakru Sirirak , He Hong , Minhas Kanwal , Foster Robert , Markert Udo R. , Pfeffer Klaus , Croy B. Anne
Published: March 2003

Gene ablation studies in mice indicate that lymphotoxin (LT)α, LTβ and LTβR are essential for the genesis of lymph nodes (LN), normal structural development of peripheral lymphoid tissues and the differentiation of natural killer (NK) cells. LTβR binds to the heterotrimeric cytokines LTα1β2 and LIGHT. LTs also regulate stromal cell expression of lymphocyte homing chemokines. Uterine decidualization in normal (+/+) mice is accompanied by the appearance and maturation of large numbers of uterine NK (uNK) cells that differentiate from precursors mobilized to the uterus from secondary lymphoid tissues. uNK cells accumulate in a transient, lymphocyte‐rich region known as the metrial gland or, more recently, the mesometrial lymphoid aggregrate of pregnancy (MLAp). To determine if LTs contribute to development of the MLAp, and to the differentiation and/or localization of uNK cells, a histological study was undertaken of implantation sites from LTα null, LTβR null and gestation day‐matched, normal mice. Implantation sites from the gene‐ablated mice contained abundant numbers of uNK cells that localized appropriately. This indicates that the stromally derived molecules supporting NK cell differentiation in the uterus differ from those used in secondary lymphoid organs.