bluuebunny/crossref_metadata_2025 huggingface dataset:
Lepromatous leprosy patients show T helper 1‐like cytokine profile with differential expression of interleukin‐10 during type 1 and 2 reactions
Authors: SREENIVASAN  , MISRA  , WILFRED  , NATH
Published: March 2003

Some leprosy patients suffer from clinical episodes associated with tissue damage which are designated as Type 1 (reversal reaction) when localized to the lesions and Type 2 (erythema nodosum leprosum, ENL) when accompanied by systemic involvement. We had reported earlier that stable, non‐reaction lepromatous leprosy subjects show T helper 2 (Th2)‐ and Th0‐ but not Th1‐like responses in the peripheral blood. To further understand the development of Th‐like responses during disease, 32 lepromatous patients undergoing reactions were studied using cytokine‐specific reverse transcription–polymerase chain reaction (RT–PCR) and enzyme‐linked immunosorbent assay (ELISA) in peripheral blood and some skin biopsies. Of interest was the evidence of a Th1‐like response with presence of interferon‐γ (IFN‐γ) and absence of interleukin‐4 (IL‐4) mRNA in the peripheral blood mononuclear cells (PBMC) of 85 and 64% of Type 1 and 2 reaction patients, respectively, and in all reaction sites. Whereas a Th0‐ was seen in some, a Th2‐like response was absent. IL‐12p40 mRNA was seen in 21/25 ENL and all Type 1 reaction subjects irrespective of the Th phenotype. IL‐12p40 and IFN‐γ were detectable in unstimulated PBMC suggesting an in vivo priming during reactions. IL‐10 was mainly associated with adherent cells and showed a differential expression in the two reactions. It was present in the PBMC of ENL but not in reversal reaction patients. Moreover, it was not detectable in the skin lesions of either type of reactions. A Th1‐like cytokine profile was associated with immunopathology and persisted up to 6–7 months after the onset of reactions.
Bradykinin stimulates interleukin‐8 production by human lung fibroblasts
Authors: HAYASHI  , YAMASHITA  , MATSUI  , MARUYAMA  , SUGIYAMA  , SUGIYAMA  , KOBAYASHI
Published: March 2003

Bradykinin (BK) is a potent inflammatory mediator that is generated from kininogens by the actions of plasma and tissue kallikreins. Lung fibroblasts have the potential to participate in the inflammatory responses by releasing proinflammatory cytokines in response to a variety of stimuli. We postulated that human lung fibroblasts might produce interleukin‐8 (IL‐8) in response to BK stimulation. The present study showed that BK stimulated human lung fibroblasts to produce IL‐8 in a dose‐ and time‐dependent manner. Furthermore, Northern blot analysis showed that BK increased IL‐8 mRNA expression. The stimulatory effect of BK on IL‐8 production was detected at the concentration of 10 nm, and the maximal stimulation was achieved with 100 to 1000 nm. Phorbol 12‐myristate 13‐acetate pretreatment diminished the ability of BK to stimulate IL‐8 production. In addition, GF109203X, a selective protein kinase C inhibitor, blocked BK‐induced IL‐8 production. These observations suggest that the stimulatory effect of BK on IL‐8 production by lung fibroblasts is, at least partially, mediated through protein kinase C. These data suggest that BK may be involved in the inflammatory reaction leading to interstitial lung disorders through stimulating IL‐8 production by lung fibroblasts.
The radiation‐attenuated schistosome vaccine induces high levels of protective immunity in the absence of B cells
Authors: ANDERSON  , COULSON  , LJUBOJEVIC  , MOUNTFORD  , WILSON
Published: March 2003

Radiation‐attenuated cercariae of Schistosoma mansoni elicit consistently high levels of protective immunity in mice. The cell‐mediated pulmonary effector mechanisms have been well characterized but the role of B cells and antibodies remains ill defined. We have compared the immune responses of B‐cell‐deficient (μMT) mice and their wild‐type (WT) counterparts following exposure to the attenuated vaccine. Both groups mounted a T helper type 1 (Th1)‐biased response in the skin‐draining lymph nodes after vaccination. Interferon‐γ was the dominant cytokine secreted by airway leucocytes after challenge in both μMT and WT mice, but there was a somewhat greater Th2 component in the former animals. The cellular infiltrates observed in the airways, and the pulmonary effector foci, were of similar composition in the two groups although some large foci were present in the μMT mice. There was a marked dichotomy in the protection induced in μMT animals by a single vaccination, with two‐thirds showing levels similar to their WT counterparts, demonstrating that cell‐mediated mechanisms alone can provide adequate protection. The remaining μMT mice had a mean worm burden identical to that of their challenge controls. A possible explanation is that a proportion of the μMT animals have a genetic defect closely associated with the μ‐heavy‐chain locus on chromosome 12, which affects their ability to mount a protective cell‐mediated response. Three vaccinations enhanced the immunity of WT animals, most likely by augmenting antibody‐mediated mechanisms. In contrast, no enhancement was seen in μMT mice, suggesting that the cell‐mediated response is not boosted by multiple exposures to attenuated larvae.
Enhancement of B7‐1 (CD80) expression on B‐lymphoma cells by irradiation
Authors: SEO  , ISHIKAWA  , NAKANO  , NAKAZAKI  , KOBAYASHI  , KAKIUCHI
Published: March 2003

Irradiation of A20.2J mouse B‐lymphoma cells enhanced their antigen‐presenting ability to induce interleukin‐2 (IL‐2) production by 42‐6A T cells specific for ovalbumin (OVA)323–339/I‐Ad. Irradiated and fixed A20.2J cells were more efficient antigen‐presenting cells (APC) to present OVA323–339 peptide than the unirradiated and fixed cells. Irradiation selectively increased the expression of B7‐1 molecules, but not of the major histocompatibility complex class II molecules, B7‐2, lymphocyte function‐associated antigen‐1, or intracellular adhesion molecule‐1. Irradiation of A20.2J cells with 100 Gy followed by overnight incubation was optimal for the enhancement of B7‐1 expression. Anti‐B7‐1 monoclonal antibody inhibited the irradiation‐induced enhancement of APC function. Irradiation of A20.2J cells induced the accumulation of B7‐1 mRNA. Thus, it was concluded that the enhancement of APC function by irradiation was due to the up‐regulation of B7‐1 molecules through the accumulation of its mRNA. Although partial inhibition of protein synthesis has been shown to enhance the accumulation of B7‐1 mRNA and its expression, irradiation did not decrease the protein synthesis in A20.2J cells. The incubation with irradiated A20.2J cells stimulated unirradiated A20.2J cells to increase B7‐1 expression, suggesting that irradiation of A20.2J cells induced expression or secretion of some molecule(s) to enhance B7‐1 expression.
Induction of the trypanosome lymphocyte‐triggering factor (TLTF) and neutralizing antibodies to the TLTF in experimental African trypanosomiasis
Authors: HAMADIEN  , LYCKE  , BAKHIET
Published: March 2003

We have demonstrated that African trypanosomes secrete a novel trypanokine, the trypanosome‐derived lymphocyte‐triggering factor (TLTF), which activates CD8+ cells to produce interferon‐γ (IFN‐γ) that in turn stimulates parasite growth. The gene for TLTF was recently cloned, and recombinant TLTF (rTLTF) showed bioactivity that was similar to native TLTF. In this work, we employed two anti‐TLTF monoclonal antibodies (mAbs) to detect levels of TLTF during Trypanosoma brucei brucei (T. b. brucei ) infections in mice. Furthermore, rTLTF was utilized to assess levels of anti‐TLTF antibodies. Mice with intact genes (wild type), and knockout mice with disrupted IFN‐γ (IFN‐γ−/−) or IFN‐γR (IFN‐γR−/−) genes were studied. The knockout mice were used in order to illustrate the role of IFN‐γ in the production of antibodies to TLTF. While wild‐type mice showed high parasitaemia accompanied by high TLTF levels and low anti‐TLTF antibodies at day 3 postinfection (p.i.), low TLTF was measured together with increased anti‐TLTF antibodies at day 21 p.i. IFN‐γ−/− mice exhibited very low parasitaemia, TLTF and anti‐TLTF antibody levels. In contrast, IFN‐γR−/− mice revealed very high parasitaemia, increased TLTF levels, but decreased anti‐TLTF antibodies. In a biological assay for TLTF, Fab′ fragments of anti‐TLTF antibodies dose dependently inhibited the TLTF‐induced IFN‐γ production by splenocytes, suggesting a regulatory importance of these antibodies. Our data demonstrate a role of IFN‐γ in the generation of neutralizing antibodies to TLTF. Furthermore, the induction of TLTF and its antibodies may constitute a new approach for future diagnosis of African trypanosomiasis.
Inhibition of T‐cell responsiveness by nasal peptide administration: influence of the thymus and differential recovery of T‐cell‐dependent functions
Authors: Metzler  , Wraith
Published: March 2003

We have previously demonstrated that intranasal (i.n.) administration of the major encephalitogenic peptide, Ac1–9 of myelin basic protein (MBP), inhibited T‐cell responsiveness in vitro and induced tolerance in the H‐2u mouse model of experimental autoimmune encephalomyelitis (EAE). The peptide analogue Ac1–9[4Y] with high‐affinity binding to the I‐Au major histocompatibility complex (MHC) class II molecule was the most effective tolerogen. Here, we show that mice pretreated with 4Y i.n. and primed with myelin had strongly reduced levels of anti‐MBP immunoglobulin G2a (IgG2a) and IgG1, demonstrating that both T helper 1 (Th1) and Th2 functions were inhibited in vivo. Since peptide administered i.n. was shown to be functionally relevant in the thymus, the time interval between 4Y i.n. and subsequent priming was varied in euthymic and adult thymectomized (ATx) mice, to examine the duration of in vitro cell unresponsiveness in the presence or absence of a thymus. For intervals of 1–6 weeks, inhibition of T‐cell proliferation was virtually complete in both euthymic and ATx mice. From 8 weeks onwards, responsiveness slowly recovered in euthymic but not in ATx mice. With an interval of 16 weeks, substantial recovery of T‐cell responsiveness in vitro in euthymic mice was reflected by a low degree of protection from EAE in vivo. By contrast, anti‐MBP IgG2a and IgG1 antibody responses were still significantly reduced. These findings suggest that T‐cell unresponsiveness by peptide i.n. represents a thymus‐independent, peripheral phenomenon, the reversal of which is confined to new T cells being exported from the thymus. As observed for EAE and antibody responses, the kinetics of recovery may vary for different effector functions.
NOS2‐derived nitric oxide regulates the size, quantity and quality of granuloma formation in Mycobacterium avium‐infected mice without affecting bacterial loads
Authors: Ehlers  , Kutsch  , Benini  , Cooper  , Hahn  , Gerdes  , Orme  , Martin  , Rietschel
Published: March 2003

Granuloma formation in response to mycobacterial infections is associated with increased expression of inducible nitric oxide synthase (NOS2) within granuloma macrophages and increased levels of nitrate/nitrite in the sera of infected mice. Continuous treatment with 5 mm or 10 mm l‐N6‐(1‐imino‐ethyl)‐lysine (L‐NIL), a selective NOS2‐inhibitor, in acidified drinking water for up to 7 weeks consistently reduced infection‐induced nitrate/nitrite to background levels in mycobacteria‐infected BALB/c mice. Oral treatment with 5 mm L‐NIL initiated at the time of infection significantly exacerbated growth of Mycobacterium tuberculosis, but had no effect on Mycobacterium avium colony‐forming unit development in the liver, spleen and lungs of intravenously infected mice. In order to examine the role of nitric oxide in mycobacteria‐induced granulomatous inflammation in the absence of any effect on the bacterial load, M. avium‐infected mice were treated with 5 mm L‐NIL from day 1 through 38 and the development of granulomatous lesions in the liver was assessed by histology, immunohistology and reverse‐transcription–polymerase chain reaction (RT‐PCR). Computer‐ and video‐assisted morphometry performed at 4 and 7 weeks post‐infection showed that treatment with L‐NIL led to markedly increased number, cellularity and size of granulomatous lesions in infected mice regardless of the virulence of the M. avium isolate used for infection. Immunohistology of the liver revealed that in mice treated with L‐NIL, the numbers of CD3+ T cells, CD21/35+ B cells, CD11b+ macrophages and RB6‐8C5+ granulocytes associated with granulomatous lesions was increased. RT‐PCR of the liver showed that in L‐NIL‐treated mice infected with M. avium, mRNA levels of tumour necrosis factor, interleukin‐12p40, interferon‐γ, interleukin‐10 and interferon‐γ‐inducible protein‐10 (IP‐10) were up‐regulated, while mRNA levels of interleukin‐4, monocyte chemotactic protein‐1 (MCP‐1) and MCP‐5 were similar to those in untreated control infected mice. When M. avium‐infected mice were treated with 5 mm L‐NIL between the 5th and 12th weeks of infection, similar changes in granuloma number and size were found in the absence of any effect on the bacterial load. These findings demonstrate that nitric oxide regulates the number, size and cellular composition of M. avium‐induced granulomas independently of antibacterial effects by modulating the cytokine profile within infected tissues.
Age‐related changes in dermal mast cell prevalence in BALB/c mice: functional importance and correlation with dermal mast cell expression of Kit
Authors: Hart P. H. , Grimbaldeston M. A. , Hosszu E. K. , Swift G. J. , Noonan F. P. , Finlay‐Jones J. J.
Published: March 2003

Differences in dermal mast cell prevalence for adult mice of different strains have been reported previously. In this study, the dermal mast cell prevalence for BALB/c and C57BL/6 mice at 6 weeks of age was similar but as BALB/c mice matured from 6 to 10 weeks of age, their dermal mast cell prevalence halved. In contrast, there was no significant difference in the dermal mast cell prevalence of 6‐ and 10‐week‐old C57BL/6 mice. These differences determined the degree of susceptibility of BALB/c and C57BL/6 mice of different ages to UVB (UV radiation of wavelength 280–320 nm)‐induced systemic immunosuppression. Expression of the receptor for stem cell factor, Kit protein, was examined on mast cells under conditions in which the dermal mast cell prevalence varied. A significant correlation was observed between Kit expression by mast cells from adult BALB/c, DBA/2 and C57BL/6 mice and dermal mast cell prevalence. In BALB/c mice, mast cell Kit expression decreased as the mice matured from 6 to 10 weeks of age and correlated with the reduction in dermal mast cell numbers. Kit levels on dermal mast cells from C57BL/6 mice were consistently higher than on mast cells from BALB/c mice although significant reductions in Kit were also measured with ageing from 6 to 10 weeks. We hypothesize that regardless of the extent of Kit expression, the dermal mast cell populations were maximally expanded in C57BL/6 mice. We suggest that BALB/c mice of 6 and 10 weeks of age are useful hosts in which to quantitatively evaluate mast cell involvement in a range of functional assays involving skin.
Simple chemicals can induce maturation and apoptosis of dendritic cells
Authors: Manome H. , Aiba S. , Tagami H.
Published: March 2003

As is well known in the case of Langerhans cells, dendritic cells (DCs) play a crucial role in the initiation of immunity to simple chemicals such as noted in the contact hypersensitivity. Because DCs are scattered in non‐lymphoid organs as immature cells, they must be activated to initiate primary antigen‐specific immune reactions. Therefore, we hypothesized that some simple chemicals must affect the function of DCs. In this paper, we first demonstrated that human monocyte‐derived DCs responded to such simple chemicals as 2,4‐dinitrochlorobenzene (DNCB), 2,4,6‐trinitrochlorobenzene (TNCB), 2,4‐dinitrofluorobenzene (DNFB), NiCl2, MnCl2, CoCl2, SnCl2, and CdSO4 by augmenting their expression of CD86 or human leucocyte antigen‐DR (HLA‐DR), down‐regulating c‐Fms expression or increasing their production of tumour necrosis factor‐α (TNF‐α). In addition, the DCs stimulated with the chemicals demonstrated increased allogeneic T‐cell stimulatory function. Next, we found that, among these chemicals, only NiCl2 and CoCl2 induced apoptosis in them. Finally, we examined the effects of these chemicals on CD86 expression by three different macrophage subsets and DCs induced from the cultures of human peripheral blood monocytes in the presence of macrophage colony‐stimulating factor (M‐CSF), M‐CSF + interleukin‐4 (IL‐4), granulocyte–macrophage colony‐stimulating factor (GM‐CSF), and GM‐CSF + IL‐4, respectively. Among them, only DCs dramatically augmented their expression of CD86. These observations have revealed unique characteristics of DCs, which convert chemical stimuli to augmentation of their antigen presenting function, although their responses to different chemicals were not necessarily uniform in the phenotypic changes, cytokine production or in the induction of apoptosis.
A mimotope peptide‐based vaccine against Schistosoma mansoni: synthesis and characterization
Authors: Arnon R. , Tarrab‐Hazdai R. , Steward M.
Published: March 2003

A panel of four mimotopes of the epitope recognized by the highly protective monoclonal antibody against Schistosoma mansoni (152‐66‐9B) was obtained by screening a solid‐phase 8mer random peptide library. Three of the four mimotopes (p28, p29 and p30) were efficiently recognized in an in vitro radioimmunoassay by the monoclonal antibody and by sera from infected mice and one (p30) induced in vitro proliferation of primed lymphocytes. When the mimotopes were conjugated to bovine serum albumin (BSA) and the conjugates used to immunize C57BL/6J mice, only the p30–BSA‐induced antibodies which were effective at complement‐mediated killing of schistosomula. The level of complement‐mediated killing obtained with the anti‐p30 antibodies was comparable to that seen with serum from mice immunized with the protective 9B‐antigen. Furthermore, following challenge infection of mimotope–BSA‐immunized mice, a greater than 40% reduction in worm burden was observed in p30–BSA‐immunized mice, a level comparable to that seen following immunization with the intact 9B‐antigen. These results show that a simple synthetic peptide immunogen comprising an eight‐amino acid mimotope of a conformational epitope on the 9B‐antigen can induce protective immune responses against S. mansoni that are comparable to those obtained following immunization with the far more complex intact antigen. This mimotope may well represent a potential component of a synthetic peptide vaccine against S. mansoni. The inclusion of other B‐cell‐ and T‐cell‐stimulating synthetic epitopes in such a vaccine, together with a more appropriate carrier, adjuvant and delivery systems may well result in a level of protection even greater than that seen with the single mimotope.
CD95 ligand expression as a mechanism of immune escape in breast cancer
Authors: Müschen M. , Moers C. , Warskulat U. , Even J. , Niederacher D. , Beckmann M. W.
Published: March 2003

Interaction of CD95 (Apo‐1/Fas) and its ligand (CD95L) plays an important role in the regulation of the immune response, since CD95+ lymphocytes may be killed after engagement of the CD95 receptor. Studying the CD95/CD95L system in 40 cases of breast cancer, the malignant cells expressed CD95L, but lost CD95 expression, when compared with non‐malignant mammary tissue. Jurkat T cells incubated on breast cancer sections underwent CD95L‐specific apoptosis. The rate of apoptosis correlated with the CD95L mRNA levels of the tissue samples. In four breast cancer cell lines, CD95L expression was increased by interferon‐γ (IFN‐γ), which resulted in higher levels of CD95L‐specific apoptosis in co‐cultured Jurkat T cells. Since IFN‐γ is mainly secreted by activated T cells, up‐regulation of CD95L in breast cancer cells in response to IFN‐γ may thus counterselect activated tumour‐infiltrating T cells and favour the immune escape of breast cancer. As demonstrated by inhibition of matrix metalloproteinases, CD95L expressed on breast cancer cells can also be shed from the cell membrane into the culture supernatant. Supernatants derived from cultured breast cancer cells induced apoptosis in Jurkat T cells via CD95L. In breast cancer patients, depletion of CD4+ and CD8+ peripheral blood lymphocytes was significantly correlated with CD95L expression in the tumours. This might be suggestive for a relationship between CD95L expression by breast cancer and systemic immunosuppression.
Forced expression of terminal deoxynucleotidyl transferase in fetal thymus resulted in a decrease in γδ T cells and random dissemination of Vγ3Vδ1 T cells in skin of newborn but not adult mice
Authors: Aono A. , Enomoto H. , Yoshida N. , Yoshizaki K. , Kishimoto T. , Komori T.
Published: March 2003

The repertoire of lymphocyte receptor genes encoded in a germline is further diversified by a number of processes, including the template‐independent addition of nucleotides (N regions) by means of terminal deoxynucleotidyl transferase (TdT). Normally, mouse γδ T cells in the early fetal thymus, whose T‐cell receptor (TCR) genes lack N regions and are encoded by Vγ3‐Jγ1 and Vδ1‐Dδ2‐Jδ2 with canonical junctions (invariant Vγ3Vδ1), are thought to be the precursors of dendritic epidermal T cells (DETC). We generated mutant mice whose endogenous TdT promoter was replaced with the lck promoter through homologous recombination. These mutant mice expressed TdT in fetal thymus, had abundant N regions and infrequent canonical junctions in γ and δ rearrangements, and showed a decreased number of γδ T cells. Various Vγ3Vδ1 T cells, most of which had N regions in their TCR genes, were found to disseminate in the skin of newborn mutant mice, whereas normal numbers of DETCs with the invariant Vγ3Vδ1 rearrangement were observed in adult mutants. These data demonstrate that the regulation of TdT expression during fetal development is important for the generation of γδ T cells, and that Vγ3Vδ1 T cells, which have various junctional sequences in their TCR genes, randomly disseminate in skin, but invariant Vγ3Vδ1 T cells have a great advantage for proliferation in skin.
Regulation of human neutrophil‐mediated cartilage proteoglycan degradation by phosphatidylinositol‐3‐kinase
Authors: Hii C. S. T. , Marin L. A. , Halliday D. , Roberton D. M. , Murray A. W. , Ferrante A.
Published: March 2003

The ability of neutrophils to degrade cartilage proteoglycan suggests that the neutrophils that accumulate in the joints of rheumatoid arthritis patients are mediators of tissue damage. The regulatory mechanisms which are relevant to the proteoglycan‐degrading activity of neutrophils are poorly understood. Since phosphatidylinositol 3‐kinase (PI3‐K), protein kinase C (PKC), the extracellular signal‐regulated protein kinase (ERK)1/ERK2 and cyclic adenosine monophosphate (cAMP) have been reported to regulate neutrophil respiratory burst and/or degranulation, a role for these signalling molecules in regulating proteoglycan degradation was investigated. Preincubation of human neutrophils with GF109203X (an inhibitor of PKC), PD98059 (an inhibitor of MEK, the upstream regulator of ERK1/ERK2) or with forskolin or dibutyryl cAMP, failed to suppress proteoglycan degradation of opsonized bovine cartilage. In contrast, preincubation of neutrophils with wortmannin or LY294002, specific inhibitors of PI3‐K, inhibited proteoglycan degradation. Incubation of neutrophils with cartilage resulted in the activation of PI3‐K in neutrophils, consistent with a role for PI3‐K in proteoglycan degradation. Activation of PI3‐K and proteoglycan degradation was enhanced by tumour necrosis factor‐α. Degradation caused by neutrophils from the synovial fluid of rheumatoid arthritis patients was also inhibited by wortmannin. These data demonstrate that the proteoglycan degradative activity of neutrophils required PI3‐K but not PKC or the ERK1/ERK2/ERK5 cascades and was insensitive to increases in intracellular cAMP concentrations.
Impaired protective immunity and T helper 2 responses in alymphoplasia (aly) mutant mice infected with Trichinella spiralis
Authors: Korenaga M. , Akimaru Y. , Shamsuzzaman S. M. , Hashiguchi Y.
Published: March 2003

The alymphoplasia (aly) mutation of mice prevents the development of systemic lymph nodes and Peyer's patches. The mutant homozygotes (aly/aly) are partially deficient in both humoral and cell‐mediated immune functions. In the present study, we show that adult worm expulsion was slightly delayed and that T helper 2 (Th2)‐type responses were partially defective in aly/aly mice after infection with Trichinella spiralis. Male aly/aly and aly/+ mice (8‐weeks old) were infected with 400 muscle larvae. There was no difference in worm recovery between the two groups on day 5. However, worm recovery in aly/aly mice was significantly higher than that in aly/+ mice on day 14. Mucosal mast cells increased in number and peaked 14 days after infection in aly/+ mice. aly/aly mice were deficient in their mucosal mast cell response through out the primary infection. To examine the existence of mast cell precursors, aly/aly mice were treated with recombinant interleukin‐3 (rIL‐3) before infection. The mast cell response was poorly induced in aly/aly mice treated with rIL‐3. An immunoglobulin E (IgE) response was not detected in aly/aly mice during the course of infection. Serum IgG1 levels in aly/aly mice were significantly lower than that of aly/+. The serum IgG2a levels increased in both strains of mice. However, IgG2a production in aly/aly mice on day 14 was half as much as that in aly/+mice. Stimulation of splenic T cells in vitro with anti‐CD3 monoclonal antibody (mAb) showed that spleen cells from aly/+ mice on day 5 produced more IL‐4 than spleen cells from aly/aly mice. IL‐4 production from aly/aly mice on day 14 was half that from aly/+ mice. Interferon‐γ (IFN‐γ) was produced in both aly/aly and aly/+ mice on day 14. Proliferation assay showed that T cells of aly/aly mice responded poorly when cultured with antigen‐presenting cells. These results suggest that aly gene is needed for the induction of protective immunity and Th2 responses in mice infected with T. spiralis.
Dendritic cells rapidly undergo apoptosis in vitro following culture with activated CD4+ Vα24 natural killer T cells expressing CD40L
Authors: Nieda M. , Kikuchi A. , Nicol A. , Koezuka Y. , Ando Y. , Ishihara S. , Lapteva N. , Yabe T. , Tokunaga K. , Tadokoro K. , Juji T.
Published: March 2003

Human Vα24 natural killer T (Vα24NKT) cells are activated by α‐glycosylceramide‐pulsed dendritic cells (DCs) in a CD1d‐dependent and T‐cell receptor‐mediated manner. There are two major subpopulations of Vα24NKT cells, CD4– CD8– Vα24NKT and CD4+ Vα24NKT cells. We have recently shown that activated CD4– CD8– Vα24NKT cells have cytotoxic activity against DCs, but knowledge of the molecules responsible for cytotoxicity of Vα24NKT cells is currently limited. We aimed to investigate whether CD4+ Vα24NKT cells also have cytotoxic activity against DCs and to determine the mechanisms underlying any observed cytotoxic activity. We demonstrated that activated CD4+ Vα24NKT cells [CD40 ligand (CD40L) ‐positive] have cytotoxic activity against DCs (strongly CD40‐positive), but not against monocytes (weakly CD40‐positive) or phytohaemagglutinin blast T cells (CD40‐negative), and that apoptosis of DCs significantly contributes to the observed cytotoxicity. The apoptosis of DCs following culture with activated CD4+ Vα24NKT cells, but not with resting CD4+ Vα24NKT cells (CD40L‐negative), was partially inhibited by anti‐CD40L mAb. Direct ligation of CD40 on the DCs by the anti‐CD40 antibody also induced apoptosis of DCs. Our results suggest that CD40–CD40L interaction plays an important role in the induction of apoptosis of DCs following culture with activated CD4+ Vα24NKT cells. The apoptosis of DCs from normal donors, triggered by the CD40–CD40L interaction, may contribute to the homeostatic regulation of the normal human immune system, preventing the interminable activation of activated CD4+ Vα24NKT cells by virtue of apoptosis of DCs.
Transforming growth factor‐β promotes ‘death by neglect’ in post‐activated human T cells
Authors: Sillett H. K. , Cruickshank S. M. , Southgate J. , Trejdosiewicz L. K.
Published: March 2003

Transforming growth factor‐β (TGF‐β) is central to the wound repair processes that follow local trauma and inflammation. In order to mimic the early events of wound‐healing, we studied the effects of TGF‐β on mitogen‐stimulated peripheral blood cells. TGF‐β added at the initiation of mitogenesis did not significantly alter T‐cell activation, proliferation, CD45 isoform switching, or activation‐induced cell death. By contrast, TGF‐β added 72 hr post‐activation (or later) enhanced the cumulative increase in apoptotic T cells. TGF‐β had no effect on mitogen‐induced up‐regulation of Fas (CD95) or Fas ligand and did not enhance killing of the Fas‐sensitive Jurkat cell line by activated T cells. Furthermore, TGF‐β had no direct effect on levels of mRNA for members of the bcl family (bcl‐X, bfl‐1, bik, bak, bax, bcl‐2 and mcl‐1). These findings suggest that TGF‐β does not directly induce apoptosis via the Fas system or by direct effects on bcl proteins. However, interleukin‐2, which can ‘rescue’ lymphocytes from spontaneous apoptosis due to cytokine deprivation, abolished the pro‐apoptotic effects of TGF‐β on post‐activated T cells, thus demonstrating that TGF‐β increases the cytokine‐dependence of T cells for survival. We propose a novel role for TGF‐β in the suppression of inflammation by promoting the elimination of post‐activated T cells once the initiating stimulus has been resolved.
The role of hepatocyte growth factor and its receptor c‐met in interactions between lymphocytes and stromal cells in secondary human lymphoid organs
Authors: Skibinski G. , Skibinska A. , James K.
Published: March 2003

Secondary lymphoid tissue consists of two major populations of cells: lymphoid cells and stromal cells. It is generally accepted that these two cell populations influence each other however, factors mediating these processes are poorly understood. In this paper we characterize one of the possible means of communication between stroma and lymphocytes namely through hepatocyte growth factor/c‐met receptor interactions. Hepatocyte growth factor (HGF) is a pleiotropic factor that is mainly produced by mesenchymal cells and acts on cells of epithelial origin which express the HGF receptor c‐met. Here we demonstrate that biologically active HGF is constitutively produced by fibroblast‐like stromal cells from human lymphoid tissues. HGF secretion from stromal cells was increased by direct contact with activated T cells. This increase was abrogated when activated T cells were separated physically from stromal cells. Using neutralizing antibody or cytokine inhibitors we provide evidence that enhancement of HGF production was due to additive effects of T‐cell membrane‐associated interleukin‐1 (IL‐1) and CD40 ligand. Finally, we also show that B lymphocytes activated with CD40L/anti‐µ or phorbol 12‐myristate 13‐acetate (PMA) express c‐met receptor. Co‐culture of activated B cells with stromal cells from spleen leads to enhanced production of immunoglobulins. This can be partially inhibited by introduction of anti‐HGF neutralizing antibodies to the culture system. Substitution of stromal cells with recombinant HGF did not produce enhancement of immunoglobulin secretion. On the other hand stimulation of c‐met receptor with HGF leads to enhanced integrin‐mediated adhesion of activated B cells to vascular cell adhesion molecule (VCAM‐1) and fibronectin. On the basis of the above experiments we conclude that HGF production by fibroblast‐like stromal cells can be modulated by activated T cells, thus providing signals for the regulation of adhesion of c‐met expressing B cells to extracellular matrix proteins. In this way HGF may indirectly influence immunoglobulin secretion by B cells.
Lipidation of T helper sequences from hepatitis C virus core significantly enhances T‐cell activity in vitro
Authors: Langhans B. , Braunschweiger I. , Schweitzer S. , Jung G. , Inchauspé G. , Sauerbruch T. , Spengler U.
Published: March 2003

Successful elimination of the hepatitis C virus (HCV) during acute infection has been linked to strong HCV‐specific in vitro T‐cell proliferation, whereas T cells from patients with chronic hepatitis C respond only weakly to HCV antigens. Lipid‐coupled peptides are immunostimulants, which might provide a basis for novel therapeutic strategies against HCV. Therefore, in 20 patients with chronic hepatitis C, we studied whether tri‐palmitoyl‐S‐cysteine‐coupled peptides could modify in vitro T‐cell proliferation (by [3H]thymidine uptake) in response to virus core and NS4. The lipopeptides corresponded to five immunodominant T helper epitopes of HCV core. Contrary to unmodified peptides, the lipopeptides specifically enhanced [3H]thymidine uptake in response to HCV antigens but not to a non‐HCV related control antigen. They increased the frequency of responders (stimulation index, SI ≥ 4) to core (13/20 versus 2/20; p = 0·0008) and NS4 (20/20 versus 7/20; p < 0·0001) among our patients with chronic hepatitis C. This immunostimulatory effect was dose‐dependent, and was observed specifically with lipopeptides corresponding to the HCV epitopes. Our data demonstrate that the poor in vitro T‐cell proliferation of patients with chronic hepatitis C can be improved when T cells are co‐stimulated with HCV core‐derived T helper lipopeptides, while the same peptides in unlipidated form had no effects. Thus, lipopeptides corresponding to HCV T‐cell epitopes may offer novel immunomodulatory strategies against HCV.
Normal and abnormal secretion by haemopoietic cells
Authors: Stinchcombe Jane C. , Griffiths Gillian M.
Published: March 2003

The secretory lysosomes found in haemopoietic cells provide a very efficient mechanism for delivering the effector proteins of many immune cells in response to antigen recognition. Although secretion shows some similarities to the secretion of specialized granules in other secretory cell types, some aspects of secretory lysosome release appear to be unique to melanocytes and cells of the haemopoietic lineage. Mast cells and platelets have provided excellent models for studying secretion, but recent advances in characterizing the immunological synapse allow a very fine dissection of the secretory process in T lymphocytes. These studies show that secretory lysosomes are secreted from the centre of the talin ring at the synapse. Proper secretion requires a series of Rab and cytoskeletal elements which play critical roles in the specialized secretion of lysosomes in haemopoietic cells.
Inhibition of the function of the FcγRIIB by a monoclonal antibody to thymic shared antigen‐1, a Ly‐6 family antigen
Authors: Ding Linna , Shevach Ethan M.
Published: March 2003

Thymic shared antigen‐1 (TSA‐1) is a member of the Ly‐6 family of glycosyl‐phosphatidylinositol (GPI)‐linked proteins. While it has been proposed that TSA‐1 may play a role in thymic development, a physiological ligand for this antigen has not been identified. Here we report that a monoclonal antibody (mAb) to TSA‐1, generated by immunizing a hamster with CD40 ligand (CD40L)‐activated B cells, interferes with the function of FcγRIIB on splenic B cells and the B‐cell lymphoma cell line, M12, by binding to TSA on the same cells. The interaction of anti‐TSA with FcγRIIB resulted in an inhibition of the ability of the FcγRIIB to cross‐link and/or aggregate soluble anti‐CD3 or soluble anti‐Cβ T‐cell receptor (TCR), leading to an inhibition of induction of expression of CD25 and CD69, interleukin (IL)‐2 production and proliferation of naive T cells. Cross‐blocking studies with mAbs strongly suggested that a physical association exists between TSA‐1 and the FcγRIIB on the surface of activated B cells and favour the view that a functional intermolecular association exists between these two distinct membrane antigens.
Requirements for autoimmune responses to mouse gastric autoantigens
Authors: Scarff Katrina L. , Judd Louise M. , Wilson Wendy E. , Laurie Karen L. , Toh Ban‐Hock , Gleeson Paul A. , Van Driel Ian R.
Published: March 2003

Autoimmune gastritis, in which the H+/K+‐ATPase of parietal cells is the major antigen, is one of the most common autoimmune diseases. Here we examined if specific properties of the H+/K+‐ATPase or parietal cells are involved in rendering them autoimmune targets. The model antigens β‐galactosidase and ovalbumin (OVA) were expressed in parietal cells of transgenic mice. On experimental induction of autoimmune gastritis by neonatal thymectomy, autoantibodies to β‐galactosidase developed in mice expressing β‐galactosidase in parietal cells, a response that was independent of either the response to the gastric H+/K+‐ATPase or gastric inflammation. In contrast, mice that expressed OVA in parietal cells did not exhibit an antibody response to OVA after thymectomy. However, increasing the frequency of anti‐OVA T lymphocytes in OVA‐expressing mice resulted in autoantibodies to OVA and gastritis. These studies indicate that parietal cells can present a variety of antigens to the immune system. Factors such as the identity and expression level of the autoantigen and the frequency of autoreactive T cells play a role in determining the prevalence and outcome of the particular immune response. In addition, as not all mice of a particular genotype displayed autoimmunity, random events are involved in determining the target of autoimmune recognition.
Infection of murine keratinocytes with herpes simplex virus type 1 induces the expression of interleukin‐10, but not interleukin‐1α or tumour necrosis factor‐α
Authors: Zak‐Prelich Malgorzata , Halliday Katrina E. , Walker Craig , Yates Catherine M. , Norval Mary , Mckenzie Roddie C.
Published: March 2003

Herpes simplex virus (HSV) is known to possess several mechanisms whereby it can evade the normal host immune defences. In this study the expression of the immunosuppressive cytokine, interleukin (IL)‐10, was monitored following infection of a murine keratinocyte cell line (PAM‐212) and compared with the expression of two proinflammatory cytokines: IL‐1α and tumour necrosis factor (TNF)‐α. The PAM‐212 cells were infected at a multiplicity of 0·5 with a clinical isolate of HSV type 1, and the mRNA of the three cytokines was assessed by semiquantitative reverse transcription–polymerase chain reaction (RT–PCR) over the following 24 hr. By 12 hr postinfection the amount of IL‐10 mRNA had increased significantly to five‐fold greater than that found in uninfected cells (P < 0·01), and this elevated level was maintained until at least 24 hr postinfection. In contrast, IL‐1α and TNF‐α mRNAs were not significantly up‐regulated by the HSV infection. Immunostaining with an IL‐10 monoclonal antibody (mAb) revealed that cytoplasmic IL‐10 protein had increased by 6–12 hr postinfection. This quantity was further increased at 24 hr postinfection, when the viral cytopathic effect was apparent. Viral replication was necessary, but not sufficient on its own, for IL‐10 induction. Experiments with HSV mutants lacking functional transactivating factors suggested that the viral transactivating proteins ICP‐0 and VP‐16 may be necessary for HSV‐induced IL‐10 expression. Thus, the up‐regulation in the expression of IL‐10 mRNA and protein induced by HSV early in the infection of keratinocytes represents a specific response and may be part of the viral strategy to avoid local immune defence mechanisms in the skin.
Nitric oxide‐enhanced resistance to oral candidiasis
Authors: Elahi Shokrollah , Pang Gerald , Ashman Robert B. , Clancy Robert
Published: March 2003

A murine model of oral candidiasis was used to show that nitric oxide (NO) is involved in host resistance to infection with Candida albicans in infection‐‘resistant’ BALB/c and infection‐‘prone’ DBA/2 mice. Following infection, increased NO production was detected in saliva. Postinfection samples of saliva inhibited the growth of yeast in vitro. Treatment with NG‐monomethyl‐l‐arginine (MMLA), an inhibitor of NO synthesis, led to reduced NO production, which correlated with an increase in C. albicans growth. Reduction in NO production following MMLA treatment correlated with an abrogation of interleukin‐4 (IL‐4), but not interferon‐γ (IFN‐γ), mRNA gene expression in regional lymph node cells. Down‐regulation of IL‐4 production was accompanied with an increase in IFN‐γ production in infection‐‘prone’ DBA/2 mice. There was a functional relationship between IL‐4 and NO production in that mice treated with anti‐IL‐4 monoclonal antibody showed a marked inhibition of NO production in saliva and in culture of cervical lymph node cells stimulated with C. albicans antigen. The results support previous conclusions that IL‐4 is associated with resistance to oral candidiasis and suggest that NO is involved in controlling colonization of the oral mucosal surface with C. albicans.
Lack of Shiga‐like toxin binding sites in germinal centres of mouse lymphoid tissues
Authors: Imai Yasuyuki , Fukui Takashi , Ikegaya Asano , Ishikawa Tomoyuki , Ono Yousuke , Kurohane Kohta
Published: March 2003

B cells in germinal centres are known to express carbohydrate antigen CD77 in human lymphoid tissues. The CD77 antigen is specifically recognized by Shiga‐like toxins (SLTs) that are produced by enterohaemorrhagic Escherichia coli O157:H7. To determine whether the binding subunits of Shiga‐like toxin‐1 (SLT‐1B) could have adverse effects on the murine immune system when used as an immunogen, we investigated whether SLT‐1B could bind to germinal centres of mouse lymphoid tissues. Frozen sections of peripheral lymph nodes and Peyer's patches from immunized mice were tested for the presence of SLT‐1B‐binding sites by immunohistological methods. Germinal centres were not stained with SLT‐1B, while they were intensely stained with peanut agglutinin (PNA), another marker of germinal centres. On the other hand, SLT‐1B specifically bound to renal tubules and collecting ducts in frozen sections of mouse kidney. This is consistent with results from human tissues. We also demonstrated that B220/PNA double‐positive populations in lymph nodes from immunized mice exhibited only marginal staining with SLT‐1B. The present results suggest that SLTs would not impede germinal centre functions of the murine immune system.
Interactions formed by individually expressed TAP1 and TAP2 polypeptide subunits
Authors: Antoniou Antony N. , Ford Stuart , Pilley Elizabeth S. , Blake Neil , Powis Simon J.
Published: March 2003

The transporter associated with antigen processing (TAP) supplies peptides into the lumen of the endoplasmic reticulum (ER) for binding by major histocompatibility complex (MHC) class I molecules. TAP comprises two polypeptides, TAP1 and TAP2, each a ‘half‐transporter’ encoding a transmembrane domain and a nucleotide‐binding domain. Immunoprecipitation of rat TAP1 and TAP2 expressed individually in the human TAP‐deficient cell line, T2, revealed that both bound the endogenously expressed HLA‐A2 and ‐B51 class I molecules. Using HLA‐encoding recombinant vaccinia viruses HLA‐A*2501, ‐B*2704, ‐B*3501 and ‐B*4402, alleles also associated with both TAP1 and TAP2. Thus, TAP1 and TAP2 do not appear to differ in their ability to interact with MHC class I alleles. Single TAP polypeptide subunits also formed MHC class I peptide‐loading complexes, and their nucleotide‐binding domains retained the ability to interact with ATP, and may permit the release of peptide‐loaded MHC class I molecules in the absence of a peptide transport cycle. It is also demonstrated by chemical cross‐linking that TAP2, but not TAP1, has the ability to form a homodimer complex both in whole cells and in detergent lysates. Together these data indicate that single TAP polypeptide subunits possess many of the features of the TAP heterodimer, demonstrating them to be useful models in the study of ATP‐binding cassette (ABC) transporters.
Host responses to Renibacterium salmoninarum and specific components of the pathogen reveal the mechanisms of immune suppression and activation
Authors: Grayson T. Hilton , Cooper Lynne F. , Wrathmell Annette B. , Evenden Janet Roper, Andrew J. , Gilpin Martyn L.
Published: March 2003

During infection, Renibacterium salmoninarum survives within the pronephric macrophages of salmonid fish. Therefore, to study the initial phases of the interaction we infected macrophages with live bacteria and analysed the responses of host and pathogen. It was found that the expression of msa encoding the p57 antigen of R. salmoninarum, was constitutive, while the expression of hly and rsh, encoding haemolysins, and lysB and grp was reduced after infection. Macrophages showed a rapid inflammatory response in which the expression of interleukin‐1β (IL‐1β), major histocompatibility complex class II (MHC II), inducible cyclo‐oxygenase (Cox‐2), and inducible nitric oxide synthase (iNOS) was enhanced, but tumour necrosis factor‐α (TNF‐α) expression was greatly reduced initially and then increased. After 5 days, except for TNF‐α and MHC II, expression returned to levels approaching those of uninfected macrophages. We propose that R. salmoninarum survives initial contact with macrophages by avoiding and/or interfering with TNF‐α‐dependent killing pathways. The effects of specific R. salmoninarum components were studied in vivo by injecting fish with DNA vaccine constructs expressing msa, hly, rsh, lysB, or grp. We found that msa reduced the expression of IL‐1β, Cox‐2, and MHC II but stimulated TNF‐α while hly, rsh and grp stimulated MHC II but down‐regulated TNF‐α. Constructs expressing hly or lysB stimulated iNOS expression and additionally, lysB stimulated TNF‐α. The results show how p57 suppresses the host immune system and suggest that the immune mechanisms for the containment of R. salmoninarum infections rely on MHC II‐ and TNF‐α‐dependent pathways. Moreover, prolonged stimulation of TNF‐α may contribute to the chronic inflammatory pathology of bacterial kidney disease.
Characterization of guinea‐pig group 1 CD1 proteins
Authors: Hiromatsu Kenji , Dascher Christopher C. , Sugita Masahiko , Gingrich‐Baker Cindy , Behar Samuel M. , LeClair Kenneth P. , Brenner Michael B. , Porcelli Steven A.
Published: March 2003

CD1 molecules are cell‐surface glycoproteins with strong structural similarities to major histocompatibility complex (MHC) class I molecules, and studies in humans and mice have demonstrated that CD1 proteins perform the unique role of presenting lipid antigens to T lymphocytes. Our previous studies have shown that guinea‐pigs, unlike the muroid rodents, have an extended family of group 1 CD1 genes. In the current study, we raised monoclonal anibodies (mAbs) against guinea‐pig CD1 proteins and generated transfected cell lines expressing individual members of the guinea‐pig CD1 family. Our results indicated that multiple members of the guinea‐pig CD1 family, including members that are homologous to the human CD1b and CD1c proteins, are expressed at the protein level in transfected cells and in specialized antigen‐presenting cells such as monocyte‐derived dendritic cells. In addition, CD1 proteins, especially guinea‐pig CD1b3, were expressed on a large number of B cells in the guinea‐pig, and CD1 expression appeared to be regulated by B‐cell maturation or differentiation. Interestingly, three different patterns of intracellular localization were observed for the various guinea‐pig CD1 isoforms, a finding that is reminiscent of the distinct patterns of intracellular localization that have been previously demonstrated for human CD1a, CD1b and CD1c. Taken together, these results provide further evidence for substantial similarities between the guinea‐pig and human CD1 systems, thus supporting the possibility that the guinea‐pig may offer significant advantages as an animal model for the study of the in vivo role of CD1 proteins in infectious and autoimmune diseases.