bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of TGF‐β on Eosinophil Chemotaxis
Authors: Luttmann  , Franz  , Matthys  , Virchow Jr
Published: March 2003

Transforming growth factor‐β (TGF‐β), which can decrease the effects of interleukin (IL)‐3, IL‐5 and granulocyte–macrophage colony‐stimulating factor (GM‐CSF) on eosinophil viability, has been shown to be chemotactic for neutrophils. However, there is little information on its effects on eosinophil chemotaxis. Because TGF‐β has recently been found in increased concentrations in asthmatic sputum, we investigated whether TGF‐β could influence eosinophil migration and eosinophil viability. Purified eosinophils from normal donors were incubated with increasing concentrations of TGF‐β. Chemotaxis was measured with a modified Boyden chamber technique. In addition, eosinophils were incubated for 96 h with either IL‐3, IL‐5 or GM‐CSF (1 ng/ml) together with increasing concentrations of TGF‐β. Eosinophil viability was then determined with propidium jodide and flowcytometry. Eosinophil chemotaxis was significantly increased in the presence of TGF‐β in concentrations between 10−9 and 10−4 μg/ml. The optimal concentration of TGF‐β in this assay was between 10−9 and 10−8 μg/ml. The chemotactic effect of TGF‐β diminished when higher as well as lower concentrations (between 10−12 and 10−3 μg/ml) were employed. In contrast, inhibition of eosinophil survival induced by IL‐3, IL‐5 and GM‐CSF reached its maximum at concentrations of TGF‐β between 10−4 and 10−3 μg/ml. From these data we conclude that TGF‐β in low concentrations can induce eosinophil chemotaxis whereas higher concentrations reduce eosinophil survival mediated by IL‐3, IL‐5 and GM‐CSF.
A Morphological Sequential Study of Mouse‐to‐Rat Cardiac Xenografts
Authors: BERSZTEL  , TUFVESON  , GANNEDAHL  , JOHNSSON
Published: March 2003

Long‐term survival of a concordant xenograft can be achieved by using cyclosporine (CyA) and deoxyspergualin (DSG) for immunosuppression. We have demonstrated in a mouse‐to‐rat heterotopic heart transplantation model that DSG treatment can be stopped after 4 weeks with the grafts remaining beating. In this investigation we have sequentially characterized the morphological changes and infiltrating cells in the transplanted hearts. Graft recipients were killed 9 days, 28 days and 56 days after transplantation. At days 9 and 28, the grafts exhibited a well‐preserved morphology, with infiltrating cells restricted only to the periphery. These cells stained positive for rat MHC class II antigens, the ED1‐macrophage marker and the CD4 antigen, and were thus considered to be macrophages. In comparison, grafts harvested at day 56 had signs of interstitial fibrosis and some arteries showed pronounced intimal thickening. There was a moderate infiltrate of cells both in the peripheral and central parts of the graft, consisting mainly of MHC class II+/CD4+/ED1+ macrophages. Very few T cells and NK cells were noticed. Termination of DSG after 28 days does not trigger a humoral rejection. However, the grafts exhibit morphological changes equivalent to those seen in chronic allograft rejection. In addition, the characteristics of the infiltrating cells conformed with cellular infiltrates associated with chronic allograft rejection. Hence, this model could in the future prove to be useful for studies of mechanisms involved in chronic xenograft rejection.
Induction of Apoptosis Reduces Immunogenicity of Human T‐Cell Lines in Mice
Authors: Ponner  , Stach  , Zoller  , Hagenhofer  , Voll  , Kalden  , Herrmann
Published: March 2003

Apoptotic cells, e.g. postinflammatory neutrophils, were reported to be engulfed by phagocytes without induction of an inflammatory response. We investigated the humoral immune response of BALB/c mice after repeated injection of viable or apoptotic human T cells. Following interleukin‐2 (IL‐2) deprivation, phytohaemagglutinin (PHA)/IL‐2 expanded human T‐cell lines were irradiated with UV‐B light to induce apoptosis, confirmed by propidium iodide staining of Triton X‐100‐lysed cells. Indirect immunofluorescence was used to detect antilymphocyte antibodies 7 days after each injection. We found high levels of antilymphocyte antibodies in all animals immunized with viable T cells, whereas animals injected with apoptotic cells showed a significantly reduced humoral immune response. We conclude that apoptotic cells induce poor xenoreactive T‐cell responses when compared with viable cells.
Immunobiological Studies on Experimental Visceral Leishmaniasis. V. The I‐ABm12 Mutation Specifies Resistance to Infection
Authors: Sen  , Roy
Published: March 2003

The I‐ABm12 mutation of the I‐Aβ subunit converted Leishmania donovani‐susceptible C57BL/6 (B6) mice into the relatively resistant B6C‐H‐2 Bm12 (Bm12) strain. The relative resistance was reflected not only in the reduced splenic and hepatic parasite burden in Bm12 (compared with B6) but also by the ability of Bm12 mice to mount a T‐cell proliferative response to parasite antigens. Assay of antileishmanial antibody (immunoglobulin G (IgG)2a and IgG1) in the sera of infected mice showed that in Bm12 mice the predominant isotype was IgG2a, rather than IgG1, whereas a similar level of both isotypes were found in B6 mice. From the serum immunoglobulin isotype titre it appeared that the antileishmanial T‐cell response was biased towards a T helper (Th) 1 response in Bm12 mice whereas it was a mixed Th1 and Th2 response in B6 mice. These observations provide credence to the notion that polymorphism in class II major histocompatibility complex (MHC) molecules is responsible for the difference in the disease phenotype.
Humoral and Cellular Immune Responses to Synthetic Peptides of the Leishmania donovani Kinetoplastid Membrane Protein‐11
Authors: Jensen  , Gasim  , Ismail  , Gaafar  , Kurtzhals  , Kemp  , El Hassan  , Kharazmi  , Theander
Published: March 2003

Native kinetoplastid membrane protein‐11 (KMP‐11), purified from crude extracts of Leishmania donovani parasites, activates T cells from individuals who have recovered from visceral leishmaniasis. In this work we used three 38‐mer peptides spanning the amino acid sequence of the L. donovani KMP‐11 as solid‐phase ligands in enzyme‐linked immunosorbent assays (ELISAs) and as stimulating antigens in lymphoproliferative assays in order to evaluate humoral and cellular immune responses to well‐defined sequences of the protein. Antibody reactivity against the three peptides was measured in plasma from 63 Sudanese visceral leishmaniasis patients (VL) and the percentage of patients with anti‐KMP‐11 antibodies in ELISA were 37% (KMP‐11‐1), 30% (KMP‐11‐2) and 58% (KMP‐11‐3). The fraction of VL patients with measurable antibody reactivity in one or more of the three ELISAs was 79%. Cross‐reactivity to the KMP‐11 peptides was detected in plasma from Sudanese patients suffering from Leishmania major infections and in plasma from Sudanese and Danish patients infected with Plasmodium falciparum. In lymphoproliferative assays, 10 of 17 PBMC isolates from donors previously infected with L. donovani showed a response to one or more of the three KMP‐11 peptides.
Neonatal Tolerance Revisited by Mathematical Modelling
Authors: Borghans  , De Boer
Published: March 2003

The classical view of a neonatal tolerance window has recently been challenged by several studies showing that neonates can evoke normal immune responses. For example, neonatal immunity against the male antigen H‐Y can be induced in female recipients by inoculation of male donor spleen cells enriched for professional antigen‐presenting cells (APCs). In the same set‐up, adult female recipients become tolerant by giving large doses of spleen cells. Using a probabilistic model, we here show how the number of T cells and endogenous APCs in the recipient, and the dose and quality of donor APCs can explain all observed phenomena. We thus reconcile the classical neonatal tolerance window with the recent data on neonatal immunity and adult tolerance.
The Three Isoforms of Transforming Growth Factor‐β Co‐stimulate Rat T Cells and Inhibit Lymphocyte Apoptosis
Authors: Schiött  , Sjögren  , Lindvall
Published: March 2003

In this study the three different mammalian isoforms of transforming growth factor‐β (TGF‐β) were compared with regard to their effect on the response of rat T lymphocytes to the superantigen, staphylococcal enterotoxin A (SEA). All different isozymes were found to increase the proliferative response of rat T lymphocytes, which was accompanied by a significantly lower percentage of apoptotic cells than proliferation in the absence of TGF‐β. The same effect of TGF‐β was observed on the generation of apoptotic cells in an allo response (mixed lymphocyte reaction). TGF‐β2 and TGF‐β3 were three to 10‐fold more potent than TGF‐β1 as co‐stimulators of T lymphocytes, and equal in decreasing the percentage of apoptotic T cells. TGF‐β1 reduced the frequency and the number of cells undergoing apoptosis in T cells and, to an even higher degree, among B lymphocytes. TGF‐β did not seem to affect the production of the apoptosis inducer, tumour necrosis factor‐α (TNF‐α), neither at the mRNA level nor at the protein level. Neutralizing antibodies against the cytokine, TNF‐α, decreased the percentage of apoptotic cells among T cells responding to SEA, both in the absence and in the presence of added TGF‐β1. Thus, when TGF‐β acts as a co‐stimulator for T‐cell activation it inhibits the induction of apoptosis and sustains the number of viable cells.
Malononitrilamides 715 and 279 Prolong Rat Cardiac Allograft Survival, Reverse Ongoing Rejection, Inhibit Allospecific Antibody Production and Interact Positively with Cyclosporin
Authors: Qi  , Ekberg
Published: March 2003

A77 1726 is a malononitrilamide (MNA) and the active metabolite of leflunomide, which has been extensively investigated and shown to be a potent immunosuppressive drug. However, the half‐life of A77 1726 is about 15–18 days in humans and leflunomide is therefore currently being developed for the treatment of autoimmune disease and not for transplantation. Search for analogues has led to the discovery of MNA 715 and 279, derivatives of A77 1726. Previous experimental experience of these compounds is still limited. The aim of the present study was to verify the efficacy of these MNAs concerning prevention and reversal of rejection, inhibition of antibody production and interaction with cyclosporin A (CsA). Heterotopic cardiac transplantation in DA to PVG rats was used. Subgroups of rats were given either CsA, MNA 715 or MNA 279 for 10 days, starting at either day 0 or day 4, or received no treatment. Titres of allospecific immunoglobulin M (IgM) and immunoglobulin G (IgG) were quantified by flow cytometry. Ten days of induction with MNA 715 or 279 produced significantly longer graft survival than in controls. Treatment from day 4 onwards, when acute rejection was established, rescued all grafts. Allospecific production of IgM or IgG was absent during MNA induction and was suppressed in animals receiving a rescue course of MNA. The transplant model was potentiated by addition of the immunomodulator quinolone‐3‐carboxamide (Linomide®), which eliminates the effect of CsA and other immunosuppressants. The combined treatment with MNA and CsA was successful in overcoming the challenge of Linomide, demonstrating the additive effects of the two drugs. In conclusion, MNA 715 and 279 were shown to be potent immunosuppressants, preventing and reversing acute allograft rejection, inhibiting and suppressing allospecific antibody production, and the drugs interacted positively with CsA.
Cellular and Cytokine Characterization of Vascular Inflammation in CBA/J Mice Chronically Infected with Trypanosoma cruzi
Authors: SUNNEMARK  , FROSTEGÅRD  , ÖRN  , HARRIS
Published: March 2003

Continuing our characterization of the immunopathological events occurring during experimental murine Chagas' disease, an immunohistological examination was conducted of the aortas of chronically infected CBA/J mice. Compared with non‐infected mice of identical age, Trypanosoma cruzi‐infected mice exhibited a marked vasculitis, with significant infiltration of inflammatory cells into the adventitial layer, including CD4+, CD8+ T cells and macrophages. Production of interleukin‐6 (IL‐6) and tumour necrosis factor‐α (TNF‐α) was evident in the inflammatory infiltrate in the endothelial and smooth muscle layers. Vasculitis was most apparent in proximity to the heart, but extended along the aorta. Such an inflammation could lead to an alteration of the endothelium, altering the protective properties of this layer and further contributing to the focal pathology characteristic of this stage of infection.
Protection Against Concanavalin A‐Induced Murine Liver Injury by the Organic Germanium Compound, Propagermanium
Authors: Ishiwata  , Yokochi  , Hashimoto  , Ninomiya  , Suzuki
Published: March 2003

Propagermanium (3‐oxygermylpropionic acid polymer) is an organic germanium compound that activates the immune system. In this study, we investigated the action of propagermanium on T‐cell‐mediated murine hepatic injury induced by concanavalin A (Con A). Oral administration of propagermanium inhibited the development of liver injury about 10 h after Con A injection. Histological analysis demonstrated that propagermanium attenuated the extent of liver damage compared with controls, reducing infiltration by leucocytes, especially CD11b‐positive cells. Infiltration by CD4‐positive cells was not affected. Tumour necrosis factor (TNF)‐α and interferon (IFN)‐γ are crucial for the development of hepatitis in this model. Propagermanium treatment induced significant inhibition of subsequent TNF‐α production about 10 h after Con A injection, without affecting IFN‐γ, interleukin (IL)‐10, IL‐4 and IL‐12 production. This effect on TNF‐α production coincided with the inhibition of aminotransferase activity late in the progression of Con A‐induced liver injury. These facts suggest that this compound affects the macrophages (Mφ) function in the liver sinusoid. Therefore, Mφ were cultured with liver sinusoidal endothelial cells (SEC) and the effect of propagermanium on TNF‐α production in the presence of IFN‐γ was determined. TNF‐α production was reduced significantly in the coculture of Mφ and SEC when Mφ was treated with propagermanium. These results might explain the mechanisms by which propagermanium inhibits Con‐A‐induced liver injury. That is, propagermanium improves hepatitis through mechanisms including the reduced production of TNF‐α, without modification of Th1‐ and Th2‐cell function.
Coordinate Up‐Regulation of the β‐Chemokine Subfamily in Autoimmune Sialoadenitis of MRL/lpr Mice
Authors: Mustafa  , Sharafeldin  , Diab  , Huang  , Bing  , Zhu  , Link  , Frithiof  , Klinge
Published: March 2003

Mononuclear cell (MNC) infiltration of the salivary and lacrimal glands is a major feature in Sjogren's syndrome (SS) and its animal model, murine autoimmune sialoadenitis (MAS). To investigate factors that influence selective infiltration by MNC of submandibular glands in young and adult MRL/lpr mice with MAS, expression of mRNA encoding the β‐chemokines monocyte chemoattractant protein (MCP)‐1, macrophage inflammatory protein (MIP)‐1α, MIP‐1β and regulated upon activation, normal T‐cell expressed and secreted (RANTES) was investigated by in situ hybridization. MCP‐1 protein production was also evaluated by immunohistochemistry. Young mice with MAS showed an early up‐regulation of mRNA expression for MCP‐1, MIP‐1β and RANTES, while MIP‐1α mRNA expression was not affected. Adult mice with MAS showed a further up‐regulation of mRNA expression for MCP‐1, MIP‐1β and RANTES, and a remarkably strong up‐regulation for MIP‐1α. Immunohistochemistry revealed that MCP‐1 protein production paralleled MCP‐1 mRNA expression in both young and adult mice. These observations implicate MCP‐1, MIP‐1β and RANTES as potential chemokines in induction of MAS, and MCP‐1, MIP‐1β, RANTES and prominently MIP‐1α in progression and perturbation of MAS.
Inhibition of HLA‐DR Expression on Activated Human Blood Eosinophils by Transforming Growth Factor‐β1
Authors: Luttmann  , Franz  , Schmidt  , Barth  , Matthys  , Virchow
Published: March 2003

Peripheral blood, bronchoalveolar lavage and sputum eosinophils of patients with asthma but not peripheral blood eosinophils from normal controls have been shown to express human leucocyte antigen (HLA)‐DR on their cell surface. Cytokines implicated in the activation of eosinophils, such as interleukin (IL)‐3 and granulocyte–macrophage colony‐stimulating factor (GM‐CSF), can up‐regulate HLA‐DR expression. However, little is known about antagonistic factors that might down‐regulate HLA‐DR expression on eosinophils. In this study we investigated whether transforming growth factor‐β (TGF‐β), which has been shown to reduce survival of activated eosinophils, can also modulate HLA‐DR expression on eosinophils. For this purpose, isolated peripheral blood eosinophils were stimulated with IL‐3 and GM‐CSF for 24 h and HLA‐DR expression was measured by flow cytometry. We found that while isolated eosinophils expressed low levels of surface HLA‐DR, incubation with GM‐CSF and IL‐3 increased HLA‐DR expression on eosinophils. TGF‐β alone did not change HLA‐DR expression on isolated eosinophils. However, co‐incubation of eosinophils with TGF‐β and either GM‐CSF or IL‐3 significantly decreased HLA‐DR expression compared to eosinophils incubated with either GM‐CSF or IL‐3 alone and this was not reversed by addition of IL‐5. This effect of TGF‐β on IL‐3‐induced HLA‐DR expression was attenuated dose‐dependently in the presence of monoclonal anti‐TGF‐β antibodies. Our results suggest that TGF‐β can reduce cytokine‐induced HLA‐DR expression on eosinophils and could thus influence eosinophil activation.
Mechanism of Regulation of Complement Receptor Type 1 Transcription by Cytosine Arabinoside in a Pre‐Erythroid Model
Authors: FUNKHOUSER  , VIK
Published: March 2003

Binding to erythrocyte complement receptor type 1 (CR1) clears immune complexes from blood and tissues, preventing complement‐mediated pathological inflammation in disease. Previous work has demonstrated that Ara‐C, a cytosine analogue, induces an 11‐fold increase in CR1 mRNA expression in K‐562 erythroleukaemia cells. In this work we therefore investigated whether the Ara‐C/K‐562 system could be used as a model for studying the pre‐erythroid regulation of CR1. We demonstrated that increased CR1 expression could be induced independently of increased haemoglobin expression. Increases in CR1 mRNA levels produced by Ara‐C treatment were not a function of increased stability of the message. However, Ara‐C induced a protein synthesis‐dependent increase in transcription initiation rate as early as 12 h after treatment. Further data suggest that the effect of Ara‐C on transcription is not a result of its direct DNA‐damaging or DNA polymerase‐inhibition activities. Induction of receptor transcription was inhibited by tyrosine kinase (TK) and protein kinase C (PKC) inhibitors. These data suggest that TK, PKC and dCTP‐adducted phospholipid signalling pathways may all play a role in the mechanism of Ara‐C‐induced CR1 transcription.
Analysis of Interleukin (IL)‐1β and Transforming Growth Factor (TGF)‐β‐Induced Signal Transduction Pathways in IL‐2 and TGF‐β Secretion and Proliferation in the Thymoma Cell Line EL4.NOB‐1
Authors: SIESE  , JAROS  , WILLIG
Published: March 2003

In the present study we investigated the interleukin (IL)‐1β and transforming growth factor‐β1 (TGF‐β1)‐mediated proliferation, and production of IL‐2 and TGF‐β, in the murine T‐cell line, EL4.NOB‐1. This cell line is resistant to TGF‐β concerning growth arrest but not autoinduction or suppression of IL‐1‐induced IL‐2 production. When cocultured with IL‐1β, TGF‐β showed growth‐promoting activity that could be antagonized by adding the phosphatidyl choline‐dependent phospholipase C (PC‐PLC) inhibitor, D609. Using specific enzyme inhibitors of protein kinases (PK) C and A, mitogen‐activated protein kinase (MAPK), phospholipase A2 (PLA2), phosphatidylinositol‐dependent (PI)‐PLC and PC‐PLC, we showed that IL‐1β‐induced IL‐2 synthesis was dependent on all investigated kinases and phospholipases, except PC‐PLC. TGF‐β1 was able to inhibit IL‐2 synthesis by the activation of PKA and MAPK. The same kinases are involved in TGF‐β autoinduction that is accompanied by a secretion of the active but not the latent growth factor and is antagonized by IL‐1β. Addition of the PI‐PLC inhibitor, ET18OCH3, or the PLA2 inhibitor (quinacrine) alone, resulted in secretion of latent TGF‐β and, in the case of ET18OCH3, active TGF‐β. These data implicate a role for PI‐PLC and PLA2 in the control of latency and secretion. Analysis of specific tyrosine activity and c‐Fos expression showed synergistic but no antagonistic effects. These events are therefore not involved in IL‐1 and TGF‐β‐regulated IL‐2 and TGF‐β production, but might participate in IL‐1/TGF‐β‐induced growth promotion.
Mechanisms Involved in Graft‐Versus‐Host Disease Induced by the Disparity of Minor Histocompatibility M1s Antigens
Authors: KAWAMURA  , KAWACHI  , KUWANO  , SUGAHARA  , WEERASINGHE  , KOSAKA,  , SEKI  , ABO
Published: March 2003

In this study we investigated which type of T cells: high T‐cell receptor (TCRhigh, cells of thymic origin) or intermediate TCR (TCRint, cells of extrathymic origin), expanded in the liver and other organs, resulting in the induction of graft‐versus‐host disease (GVHD) with minor lymphocyte stimulating (M1s) disparity. When 6.5 Gy‐irradiated BALB/c (H‐2d M1s‐1b2a) mice were injected with interleukin‐2 receptor β‐chain− (IL‐2Rβ−) CD3high cells purified from the spleen of B10.D2 (H‐2d M1s‐1b2b) mice, IL‐2Rβ+CD3high cells expanded in the liver and other organs of recipient mice. The majority of these cells were found to be IL‐2Rα−Mel‐14−CD4+Vβ3+ in GVHD mice. The CDR3 region in their TCR‐αβ (i.e. N–Dβ–N) was polyclonal, although there were skewed usages of Vβ3 and Jβ2.4. The majority of cells were confirmed to be of donor origin by the individual discrimination method, namely, they originated from isolated IL‐2Rβ−CD3high cells. Interestingly, these T cells lacked cytotoxicity against both a natural killer (NK)‐sensitive target and thymocytes with M1s disparity and nondisparity. Another important finding was that activated granulocytes expanded at generalized sites in GVHD mice. The present results raise the possibility that M1s disparity is mainly recognized by TCRhigh cells with unique properties but that direct effector cells that induce GVHD might not be such T cells but rather accompanied granulocytes.
Localization of Interleukin‐13 Gene‐Expressing Cells in Tuberculin Reactions and Lesional Skin from Patients with Atopic Dermatitis
Authors: VAN DER PLOEG  , MATUSEVICIENE  , FRANSSON  , WAHLGREN  , OLSSON  , SCHEYNIUS
Published: March 2003

Interleukin (IL)‐13 is a cytokine thought to play an important role in IgE‐mediated allergic reactions. The aims of the present study were to localize IL‐13 mRNA in positive tuberculin reactions and atopic dermatitis lesions using in situ hybridization and to study the possible influence of atopy on the cytokine gene expression in tuberculin reactions. Punch biopsy specimens were taken from tuberculin reactions in 17 nonatopic and 12 atopic (Phadiatop positive), but otherwise healthy, individuals 72 h after injection of purified protein derivative, from chronic lichenified lesions and nonlesional skin in six patients with atopic dermatitis and from normal skin in 12 healthy individuals. IL‐13‐mRNA‐producing cells were located mainly in the dermal‐cell infiltrates, both in atopic dermatitis lesions and tuberculin reactions. In the atopic dermatitis lesions, IL‐13‐mRNA expression was found in close apposition to the epidermis. Higher numbers of IL‐13‐mRNA‐producing cells were observed in the dermal‐cell infiltrates of chronic lichenified skin lesions of patients with atopic dermatitis (13.3 cells/mm2, range 0.6–42.4 cells/mm2) than in the tuberculin reactions (1.1 cells/mm2, range 0–3.8 cells/mm2) (P < 0.01) despite the larger cell infiltrates in the tuberculin reactions. No significant difference in IL‐13 or interferon‐γ gene expression in the tuberculin reactions was seen between atopic and nonatopic individuals.
Characterization of Antibody Responses to a Plasmodium falciparum Blood‐Stage Antigen Induced by a DNA Prime/Protein Boost Immunization Protocol
Authors: Haddad  , Liljeqvist  , StÅhl  , Hansson  , Perlmann  , Ahlborg  , Berzins
Published: March 2003

The humoral immune responses elicited by priming with a DNA plasmid and boosting with either the plasmid or the corresponding recombinant protein in alum adjuvant were compared. The plasmid DNA encoded a sequence (M3) derived from the Plasmodium falciparum antigen Pf155/RESA, and the recombinant protein consisted of the identical malarial sequence fused to an albumin‐binding region (BB) of streptococcal protein G. Mice of different genetic backgrounds (CBA, Balb/c and C57Bl/6) were primed with plasmid DNA and boosted with either plasmid or recombinant protein. In all strains of mice, boosting with protein elicited higher anti‐M3 antibody levels than obtained by boosting with plasmid, yet the kinetics and longevity of the secondary responses were comparable. Antiserum obtained after protein boosting displayed an immunoglobulin (Ig)G subclass profile skewed to the IgG1 isotype, regardless of the mouse strain. In contrast, mice receiving a second injection with plasmid responded with a more mixed IgG subclass profile. Inclusion of a P. falciparum circumsporozoite protein‐derived T‐helper epitope (CS.T3) in the immunization plasmid as well as in the fusion protein, did not significantly change the humoral responses to M3. The results show the potential of DNA vaccination for the purpose of priming an antibody response against the malarial blood‐stage antigen Pf155/RESA. When combined with a protein boost, this DNA priming results in high‐titred and long‐lasting anamnestic responses.
Interaction Between B.7 and CD28 Costimulatory Molecules is Essential for the Activation of Effector Function Mediating Spontaneous Tumour Regression
Authors: RAO  , VARALAKSHMI CH. , KUMARI  , KHAR
Published: March 2003

The spontaneous regression of a rat histiocytoma, AK‐5, is mediated by activated natural killer cells through antibody‐dependent cellular cytotoxicity. In addition to the Fc–FcR interaction between the target and the effector cells demonstrated previously, we show the participation of costimulatory molecules B7 and CD28 in the efficient killing of the tumour cell. Blockade of the costimulatory interaction in vivo using anti‐CD28 led to increased tumour growth and a suppressed cytokine response. Anti‐CD28 antibody administration in vivo also diminished the cytotoxic potential of NK cells against AK‐5 cells in vitro. Our studies also demonstrate the expression of B7.1 and B7.2 antigen on AK‐5 tumour cells. The cytotoxic activity of natural killer cells was significantly inhibited when the effector/target cells were cultured in the presence of antibodies raised against B7.1, B7.2 and CD28. Administration of anti‐CD28 in vivo also affected the efficiency of the formation of effector/target conjugates in vitro. Similarly, anti‐CD28 injections affected expression of the adhesion molecules LFA 1 and ICAM 1 by splenocytes. Administration of anti‐B7.1 and B7.2 antibodies in AK‐5 tumour‐bearing animals showed a differential response. The cytotoxicity of natural killer cells was significantly inhibited after anti‐B7.2 administration, suggesting the preferential participation of B7.2 molecules in vivo. These observations suggest an important role for B7–CD28 interaction in AK‐5 tumour regression.
Direct Binding of Purified HLA Class I Antigens by Soluble NKG2/CD94 C‐Type Lectins from Natural Killer Cells
Authors: Ding  , Sumitran  , Holgersson
Published: March 2003

Human natural killer (NK) cell cytotoxicity is inhibited following human leucocyte antigen (HLA) class I binding by killer cell inhibitory receptors belonging to the immunoglobulin or C‐type lectin protein families. Of the latter family, CD94 and NKG2A or ‐B associate to inhibit NK cell cytotoxicity. We have constructed C‐Myc epitope‐tagged soluble NKG2A, ‐B, ‐C, ‐D and human NKR‐P1 lectin domains, and studied their ability to associate with Flu‐tagged soluble CD94 lectin domains. Furthermore, their ability to bind solubilized immunoaffinity‐purified HLA class I antigens, either alone or following association with CD94 lectin domains, was evaluated using flow cytometry and Western blot analyses. We show that soluble NKG2A, ‐B and ‐C lectin domains interact with CD94 lectin domains to form complexes, whereas NKG2D and human NKR‐P1 lectin domains do not. Soluble NKG2C, ‐D and CD94 lectin domains bind solubilized affinity purified HLA class I antigens on their own, whereas NKG2A and ‐B require association with CD94 lectin domains for binding. Soluble human NKR‐P1 lectin domains do not bind solubilized HLA class I antigens in our system.
Testicular‐Associated Immune Deviation and Prevention of Adjuvant‐Induced Arthritis by Three Tolerization Methods
Authors: Ditzian‐Kadanoff
Published: March 2003

A new tolerizing effect, testicular‐associated immune deviation (TAID), was produced in rats by injecting antigen, emulsified with adjuvant, into the testicle. TAID abrogated the delayed hypersensitivity response to subsequent immunization. This study also assessed the effect of anterior chamber‐associated immune deviation (ACAID), the aforementioned TAID and an equivalent tolerization method named alternative in vitro immune deviation (AVID), on adjuvant‐induced arthritis (AIA). ACAID was produced by introducing antigen, emulsified with adjuvant, into the anterior chamber of the eye. AVID was achieved by exposing peritoneal exudate cells to antigen in the presence of fetal calf serum in vitro and then injecting the cells intraperitoneally. Antigen in adjuvant was administered intradermally to Dark Agouti rats, which are normally susceptible to AIA. One week after treatment with ACAID, TAID or AVID, the rats became resistant to AIA. The presence of neutralizing antibody to transforming growth factor‐β2 (TGF‐β2) in the culture abrogated the AVID effect. We concluded that introducing an antigen to the testicle induces immune deviation, and that prior introduction of the antigen to macrophages in an appropriate immune suppressive context, such as ACAID, TAID or AVID, prevents AIA.
Influence of Immunopotentiators on the Antiporin Immunoglobulin G Subclass: Distribution and Protective Immunity Against Murine Salmonellosis
Authors: Nandakumar  , Muthukkaruppan
Published: March 2003

To improve the immune potential of porin (a pore‐forming protein of Salmonella sp.), different immunopotentiators such as Freund's complete adjuvant (FCA), lipopolysaccharide (LPS) and polyoxydonium (PO) were evaluated by studying the nature of the protective immune response induced against murine Salmonellosis. The nontoxic, synthetic heteropolymer polyoxydonium was as good as LPS at inducing antiporin immunoglobulin G (IgG) antibodies and protective immunity. Analysis of the antiporin IgG subclass pattern revealed a preferential increase in a particular subclass based on the immunopotentiator used. Porin, alone or emulsified in FCA, elicited predominantly antiporin IgG1 antibodies, whereas LPS preferentially evoked antiporin IgG2a, IgG2b and IgG3 antibodies. Polyoxydonium induced a clear shift towards antiporin IgG2b antibodies. The significance of these antiporin IgG subclass antibodies in protection against murine Salmonellosis was studied by passive immunization and by analysing the infected mouse sera.
Low‐Dose Mercuric Chloride Induces Resistance in Brown Norway Rats to Further Mercuric Chloride by Up‐Regulation of Interferon‐γ
Authors: Szeto  , Gillespie  , Mathieson
Published: March 2003

Mercuric chloride induces autoimmunity in Brown Norway rats with polyclonal B‐cell activation, hyper‐IgE and multiple autoantibodies. Pre‐treatment with low‐dose HgCl2 (one‐tenth of the standard dose) induces resistance to later full‐dose HgCl2; we have studied the mechanism of this resistance. Brown Norway rats given low‐dose HgCl2 showed only a modest increase in serum IgE level, three logs lower than rats given standard‐dose HgCl2, and no up‐regulation of splenic interleukin (IL)‐4 mRNA. There was up‐regulation of splenic interferon (IFN)‐γ gene expression and a progressive rise in serum IFN‐γ. Neither IL‐12 nor IL‐18 were induced, but there was up‐regulation of IL‐12 receptor β2‐chain (IL‐12Rβ2) expression. IL‐10 and transforming growth factor (TGF)‐β expression did not change. Serum IgE and splenic IL‐4 mRNA expression remained static when these rats were rechallenged, confirming resistance. Thereafter IFN‐γ expression gradually fell, after which IL‐4 expression and serum IgE rose slightly. Our observations suggest that low‐dose HgCl2 confers protection in Brown Norway rats to further HgCl2 by up‐regulation of IFN‐γ, associated with enhanced IL‐12Rβ2 expression. The immunological response to HgCl2 in susceptible rat strains is more complex than previously appreciated and is dose dependent, with low doses inducing a T helper ‘(Th)1’ type of response in contrast to the ‘Th2’ type response associated with standard doses.
Lactic Dehydrogenase Virus Inhibits Allergic Immunoglobulin E Production: In Vivo Molecular Analysis of Cytokines
Authors: Morimoto  , Iwata  , Hayashi
Published: March 2003

The effects of lactic dehydrogenase virus (LDV) infection on allergic immunoglobulin (Ig)E production and interleukin (IL)‐4 gene expression were studied. LDV infection suppressed antigen‐induced IgE production in sensitized mice. The elevations of IL‐4 gene expression in spleen and mesenteric lymph nodes 3 and 7 days after ovalbumin challenge were suppressed significantly in LDV‐infected mice compared with control mice. The expression of the interferon (IFN)‐γ gene of mesenteric lymph nodes was significantly increased in LDV‐infected mice. These results suggest that LDV infection suppressed antigen‐induced IgE production by decreasing IL‐4 production, and that suppression of IL‐4 gene expression may be mediated by a mutual inhibition mechanism between T helper (Th)1 and Th2 cells.
Interleukin‐15 in Mycobacterial Infection of Antigen‐Presenting Cells
Authors: Maeurer  , Seliger  , Trinder  , Gerdes  , Seitzer
Published: March 2003

Interleukin‐15 (IL‐15) shares many biological functions with IL‐2 but also exhibits unique effects. Some of these represent the potent chemoattractant activity and expansion of distinct T‐cell subsets, particularly memory T cells. IL‐15 may therefore modulate the quality and quantity of cellular immune responses directed against intracellular pathogens. Immunohistochemical examination of skin lesions obtained from patients with the lepromatous or the tuberculoid form of Hansen's disease revealed intralesional IL‐15 protein in both forms of the disease. In addition to Mycobacterium leprae, a number of different mycobacterial species are capable of effectively inducing IL‐15 secretion in infected macrophages. In this work, increased IL‐15 secretion was observed in IL‐4/granulocyte–macrophage colony‐stimulating factor (GM‐CSF)‐activated antigen‐presenting cells (APC) compared with unstimulated macrophages. Immunocytological detection of intracellular IL‐15 revealed that infection with different mycobacterial species resulted in different staining patterns of anti‐IL‐15 immunoreactive material in APC. In contrast to IL‐2 or IL‐7, IL‐15 enhanced the cytolytic potential of immune effector cells in vitro and favoured the expansion of CD1b‐restricted immune cells recognizing mycobacterial‐associated antigens presented by autologous APC. IL‐15 produced by infected cells in situ may represent one of the key cytokines involved in granuloma formation and may aid the augmentation of cellular immune responses directed against mycobacterial‐infected cells.
Participation of CD1 Molecules in the Presentation of Bacterial Protein Antigens in Humans
Authors: Ulanova  , Tarkowski  , Hahn‐Zoric  , Hanson
Published: March 2003

Human CD1 molecules, expressed on the surface of professional antigen‐presenting cells (including dendritic cells, Langerhans' cells, B cells and activated monocytes) are structurally homologous to major histocompatibility complex (MHC) class I and class II molecules. CD1b and CD1c have been shown to present nonpeptide bacterial antigens to T cells. We hypothesized that CD1 molecules may also be involved in the presentation of bacterial protein antigens. Human peripheral blood mononuclear cells (PBMC) were exposed to two medically important proteins, tetanus toxoid (TT) and purified protein derivative (PPD), with and without murine monoclonal antibodies (MoAbs) specific for CD1a, CD1b and CD1c. All the MoAbs substantially inhibited the proliferative responses of PBMC to TT and PPD. Simultaneous interaction of CD1 and MHC class II molecules was even more inhibitory to these antigen‐specific proliferative responses. In contrast, neither mixed lymphocyte reaction nor superantigen and mitogenic responses were affected by CD1‐specific antibodies, indicating a certain restriction pattern in antigen presentation. Our findings suggest that, besides MHC class I and II molecules, there is a family of nonpolymorphic cell surface molecules that is able to present certain bacterial protein antigens to T cells.
Minute Defects in the Expression of MHC E Molecules Lead to Impaired Protection from Autoimmunity in NOD Mice
Authors: Rietz  , Pilström  , Brenden  , Böhme
Published: March 2003

The E complex of the major histocompatibility complex (MHC) can prevent the spontaneous development of diabetes in nonobese diabetic (NOD) mice transgenic for the Ea gene. None of three promoter‐mutated Ea constructs with Ea expression directed to different subsets of immunocompetent cells exerts full protection in NOD mice. The promoter‐mutated constructs are all capable of mediating intrathymic elimination of I‐E‐restricted T cells. Thus, thymic negative selection is not responsible for the protective effect but a more complex effect is likely. Here we show that combinations of two or three different mutated Ea constructs do not protect against intra‐islet insulitis either. We also show that spleen cells from protected animals are sufficient to protect NOD mice in adoptive transfer experiments. The only detectable expression defects in splenic cells or cells influencing the repertoire of splenic cells are in the B‐cell compartment. Furthermore, in three construct combinations, the differences to wild‐type expression are extremely small. Thus, we conclude that even minute disturbances of the E expression pattern might reduce the protection of NOD mice from insulitis.
Reduction in Interleukin‐2‐Producing Cells but not Th1 to Th2 Shift in Moderate and Advanced Stages of Human Immunodeficiency Virus Type‐1‐Infection: Direct Analysis of Intracellular Cytokine Concentrations in CD4+ CD8− T Cells
Authors: Tanaka  , Hirabayashi  , Gatanaga  , Aizawa  , Hachiya  , Takahashi  , Tashiro  , Kohsaka  , Oyamada  , Ida  , Oka
Published: March 2003

Previous studies have suggested that CD4+ T lymphocytes shift from the Th1 type to the Th2 type during disease progression in patients with the human immunodeficiency virus type‐1 (HIV‐1). In the present study, we used a modified method that allowed a direct measurement of intracellular cytokines in CD4+ CD8− T cells. A total of 48 HIV‐1‐infected (HIV+) and 16 HIV‐1‐uninfected (HIV‐) individuals were studied. The percentages of CD4+ CD8− T cells producing interleukin‐2 (IL‐2), interferon‐γ (IFN‐γ), interleukin‐4 (IL‐4), or interleukin‐5 (IL‐5) in HIV+ and HIV‐ subjects were 23.6% versus 34.9% (P < 0.01), 13.7% versus 13.2%, 1.3% versus 1.0%, and 1.2% versus 0.9%, respectively. The population of IL‐2‐producing cells decreased proportionately with reductions in CD4 counts (< 200/mm3, 200–500/mm3, and > 500/mm3 to 18.0%, 23.5%, and 30.5%, P < 0.05, respectively). There was an inverse correlation between the percentage of IL‐2‐producing cells and plasma viral load (r = − 0.446, P < 0.05). However, the percentages of CD4+ CD8− T cells producing other cytokines were not different between HIV+ and HIV−. Our cross‐sectional study demonstrated a decrease in IL‐2‐producing cells but not the Th1 to the Th2 shift in the CD4+ CD8− T cell population in the moderate and advanced stages of HIV‐1‐infection.
Beta1‐Integrins Control Spontaneous Adhesion and Motility of Human Progenitor Thymocytes and Regulate Differentiation‐Dependent Expression of the Receptor for Hyaluronan‐Mediated Motility
Authors: Gares  , Pilarski
Published: March 2003

The functions of the receptor for hyaluronan‐mediated motility (RHAMM) and β1‐integrin in adhesion and motility were analysed for human progenitor multinegative (CD3− 4− 8− 19−) thymocytes (MN Thy). Both α4β1‐ and α5β1‐integrins are expressed by MN Thy, but only α4β1 mediates fibronectin (FN)‐dependent adhesion and motility. Freshly isolated MN Thy lack expression of RHAMM and their motility is RHAMM independent. Prolonged surface expression of RHAMM on MN Thy is dependent upon FN. RHAMM expression, which occurs prior to surface expression of CD3/T‐cell receptor (TCR), was found to be inhibited by cross‐linking of α4‐, α5‐ and β1‐integrins, as was the prolonged FN‐dependent phase of RHAMM expression. To confirm that RHAMM expression had been down‐regulated rather than rendered cryptic by treatment with immobilized anti‐integrin monoclonal antibody (MoAb), RHAMM mRNA levels were analysed. Transcription of RHAMM was decreased 7–12‐fold by treatment with immobilized anti‐α4 or anti‐α5, and twofold by anti‐β1. Prior to expression of CD3/TCR and RHAMM, α4β1 regulates migratory behaviour. After MN Thy differentiate to acquire CD3/TCR in vitro or in vivo, their motility becomes dependent upon both RHAMM and β1‐integrins. Integrins play a direct role in FN‐dependent, RHAMM‐independent motility of MN Thy, and an indirect role in RHAMM‐dependent motility. This work shows that β1‐integrins are primary mediators and regulators of fundamental cell behaviours required during migratory phases of T‐cell differentiation that occur prior to the expression of CD3/TCR.
Genetics of Inflammatory Bowel Disease: A Reappraisal
Authors: Orchard  , Satsangi  , Van Heel  , Jewell
Published: March 2003

The advent of advanced molecular biological techniques in the last two decades has allowed the study of genetic factors in inflammatory bowel disease (IBD). A variety of techniques have been employed to elucidate the effects of genes, starting with the clinical observations that IBD is more common in the relatives of patients than the general population, and the consistency of clinical features within families. The situation is likely to be much more complicated than single gene disorders, and it is estimated that between 10 and 20 genes may be involved. Genome scanning techniques using microsatellite markers have been employed to highlight areas of chromosomes linked to disease such as those on chromosomes 12 and 16. In addition association studies of specific genes such as HLA and cytokine genes have been carried out on functional or positional grounds. It is likely that a combination of these techniques will be required to elucidate the role of individual genes. Recently much work has been focused on genes that may determine clinical phenotype such as disease extent or severity or the response to treatment. Identification of these genes may lead to better targeting of therapy and prognostication, and they are likely to be easier to identify than disease susceptibility genes.
Identification and Linkage of the Proteasome Activator Complex PA28 Subunit Genes in Zebrafish
Authors: Murray  , Sültmann  , Klein
Published: March 2003

PA28 is an activator of the latent 20S proteasome, a large multisubunit complex involved in intracellular proteolysis. Two forms of hexameric PA28 have been identified, PA28‐(αβ)3 and PA28‐(γ)6, of which the former is of immunological importance. Both the PA28‐α and PA28‐β subunits are inducible by interferon‐γ (IFN‐γ) and the PA28‐(αβ)3 complex enhances the ability of the 20S proteasome to produce peptides suited for binding to major histocompatibility complex (Mhc) class I molecules. To identify the homologues of the PA28 subunits in zebrafish we screened a cDNA library and obtained full‐length cDNA sequences of the genes PSME1, PSME2 and PSME3 coding for the PA28‐α, PA28‐β and PA28‐γ subunits, respectively. Phylogenetic analysis indicates the existence of the ancestors of all three genes prior to the divergence of tetrapods and bony fishes. The IFN‐γ‐inducible subunits, PA28‐α and PA28‐β, evolve faster than the presumably older PA28‐γ subunit. Using zebrafish radiation hybrid panels, the genes PSME2 and PSME3 were mapped to linkage group 12 and shown to be separated by a distance of less than 2.4 cm. This observation suggests that an intrachromosomal duplication event created the precursor of the IFN‐γ‐inducible genes from a PA28‐γ‐like ancestor prior to their recruitment into the Mhc class I peptide presentation pathway.
HLA‐A, B, Cw and DRB1, DRB3/4/5, DQB1, DPB1 Frequencies in German Immunoglobulin A‐Deficient Individuals
Authors: Machulla  , Schönermarck  , Schaaf  , Müller  , Kloß  , Krüger  , Kunze  , Schönermarck  , Langner
Published: March 2003

HLA class I and II frequencies and haplotype frequencies were determined in 80 German immunoglobulin (Ig)A‐deficient individuals and 157 healthy controls with normal IgA levels using serological and DNA typing methods. For several alleles, significant associations were found, which could be explained mainly in the context of a positive association with three different extended haplotypes (HLA‐B*08:DRB1*0301: DQB1*0201, HLA‐B*14:DRB1*0102:DQB1*0501 and HLA‐B*44:DRB1*0701:DQB1*0202) and a negative association with a fourth haplotype (HLA‐B*07:DRB1*1501:DQB1*0602). Furthermore, for the first time this study reports a positive association of IgA deficiency with DPB1 alleles. Homozygosity rate for the gene loci DRB1 and DQB1 was increased in IgA deficiency. Further analysis suggested a different pattern of HLA associations depending on the degree of IgA deficiency and the gender of the IgA‐deficient individuals.
Maternal Antibodies to HIV‐1 Envelope Domains: No Correlation with HIV‐1 Vertical Transmission in Patients from Argentina
Authors: Calarota  , Libonatti
Published: March 2003

The role of the maternal antibody response in relation to vertical human immunodeficiency virus type 1 (HIV‐1) transmission was investigated in HIV‐1‐infected mothers from Argentina. Sera from 23 transmitting and 18 nontransmitting HIV‐1‐infected mothers were tested for the presence of antibodies to V3 loop gp120 peptides representing both Argentinian sequences and several well‐characterized viral isolates from different geographic areas. Argentinian sera from transmitting mothers had significantly higher capacity to react with four of 14 V3 loop peptides tested than sera from nontransmitting mothers. Frequency of reactivity against the other peptides did not differ between the two maternal groups. Furthermore, no differences in antibody affinity were found between transmitting and nontransmitting mothers. Sera were also tested against overlapping peptides covering a neutralizing epitope of the HIV‐1 MN gp41 (amino acids 648–677). Statistical analysis indicated that no correlation between anti‐gp41 antibodies and vertical transmission exists. Although we used V3 loop peptides based on local HIV‐1 sequences, our data showed that maternal antibodies to these peptides, as well as to gp41 peptides, are not correlated with protection against HIV‐1 vertical transmission.