bluuebunny/crossref_metadata_2025 huggingface dataset:
Psychopharmacological treatment with lithium and antiepileptic drugs: suggested guidelines from the Danish Psychiatric Association and the Child and Adolescent Psychiatric Association in Denmark
Authors: Licht R. W. , Vestergaard P. , Kessing L. V. , Larsen J. K. , Thomsen P. H.
Published: September 2003

A subcommittee under the Danish Psychiatric Association and the Child and Adolescent Psychiatric Association in Denmark have recently developed national guidelines for the psychopharmacological treatment with lithium and antiepileptic drugs, and the present translation aims at contributing to the international discussion on the development of proper guidelines for the treatment of bipolar disorder. Among the antiepileptic drugs, the report deals with valproate, carbamazepine and lamotrigine and to a lesser extent with oxcarbazepine, gabapentin and topiramate. The various drugs will be reviewed, outlining the scientific evidence for mood‐stabilizing properties and discussing major side effects, the most important interactions with other drugs and practical use. Special considerations during pregnancy and lactation, during treatment of children and adolescents and during treatment of the elderly will also be presented. Antidepressants and antipsychotics are beyond the scope of the report, but due to the mood‐stabilizing properties of at least some of the atypical antipsychotics, these agents will be brought into some focus in connection with the overall treatment guidelines for the different phases of bipolar disorder given at the end of this report.
Some interfaces of dendritic cell biology
Authors: STEINMAN RALPH M.
Published: September 2003

The field of dendritic cell (DC) biology is robust, with several new approaches to analyze their role in vivo and many newly recognized functions in the control of immunity and tolerance. There also is no shortage of mysteries and challenges. To introduce this volume, I would like to summarize four interfaces of DC research with other lines of investigation and highlight some current issues. One interface is with hematopoiesis. DCs constitute a distinct lineage of white blood cell development with some unique features, such as their origin from both lymphoid and myeloid progenitors, the existence of several distinct subsets, and an important final stage of differentiation termed “maturation,” which occurs in response to inflammation and infection, and is pivotal for determining the subsequent immune response. A second interface is with lymphocyte biology. DCs are now known to influence many different classes of lymphocytes (B, NK, NKT) and many types of T cell responses (Th1/Th2, regulatory T cells, peripheral T cell deletion), not just the initial priming or induction of T cell‐mediated immunity, which was the first function to be uncovered. DCs are sentinels, controlling many of the afferent or inductive limbs of immune function, alerting the immune system and controlling its early decisions. A third interface is with cell biology. This is a critical discipline to understand at the subcellular and molecular levels the distinct capacities of DCs to handle antigens, to move about the body in a directed way, to bind and activate lymphocytes, and to exert many quality controls on the type of responses, for both tolerance and immunity. A fourth interface is with medicine. Here DCs are providing new approaches to disease pathogenesis and therapy. This interface is perhaps the most demanding, because it requires research with humans. Human research currently is being slowed by the need to deal with many challenges in the design of such studies, and the need to excite, attract and support the young scientists who are essential to move human investigation forward. Nonetheless, DCs are providing new opportunities to study patients and the many clinical conditions that involve the immune system.
Pathogens target DC‐SIGN to influence their fate DC‐SIGN functions as a pathogen receptor with broad specificity
Authors: GEIJTENBEEK TEUNIS B. H. , VAN KOOYK YVETTE
Published: September 2003

Dendritic cells (DC) are vital in the defense against pathogens. To sense pathogens DC express pathogen recognition receptors such as toll‐like receptors (TLR) and C‐type lectins that recognize different fragments of pathogens, and subsequently activate or present pathogen fragments to T cells. It is now becoming evident that some pathogens subvert DC functions to escape immune surveillance. HIV‐1 targets the DC‐specific C‐type lectin DC‐SIGN to hijack DC for viral dissemination. HIV‐1 binding to DC‐SIGN protects HIV‐1 from antigen processing and facilitates its transport to lymphoid tissues, where DC‐SIGN promotes HIV‐1 infection of T cells. Recent studies demonstrate that DC‐SIGN is a more universal pathogen receptor that also recognizes Ebola, cytomegalovirus and mycobacteria. Mycobacterium tuberculosis targets DC‐SIGN by a mechanism that is distinct from that of HIV‐1, leading to inhibition of the immunostimulatory function of DC and pathogen survival. Thus, a better understanding of DC‐SIGN‐pathogen interactions and their effects on DC function is necessary to combat infections.
Dendritic cells as inducers of antimicrobial immunity in vivo
Authors: SUNDQUIST MALIN , JOHANSSON CECILIA , WICK MARY JO
Published: September 2003

Models of infection have provided important insight into the function of dendritic cells (DC) in vivo. Several microbial products induce DC maturation via Toll‐like receptors, a process that is crucial for the ability of DC to initiate adaptive immune responses. Splenic DC have also been shown to produce IL‐12 during infection in vivo. This DC‐derived IL‐12 might be important to skew T cell responses towards Th1. Microbial infections also induce changes in the DC populations of lymphoid organs, often in a subset‐specific manner, manifested as an accumulation and redistribution of DC. Furthermore, data are emerging pointing at an absolute requirement of DC in priming of naïve T cells in vivo.
Langerhans cells – dendritic cells of the epidermis
Authors: ROMANI NIKOLAUS , HOLZMANN SANDRA , TRIPP CHRISTOPH H. , KOCH FRANZ , STOITZNER PATRIZIA
Published: September 2003

Langerhans cells (LC) are dendritic cells of the epidermis. They are highly specialized leukocytes that serve immunogenic and tolerogenic purposes. Here, we review some aspects of LC biology, emphasizing those areas where LC are or may turn out to be special.
Airway dendritic cells: Co‐ordinators of immunological homeostasis and immunity in the respiratory tract
Authors: STUMBLES PHILIP A. , UPHAM JOHN W. , HOLT PATRICK G.
Published: September 2003

The large quantities and complex mixtures of antigens encountered daily at airway mucosal and alveolar surfaces pose a major challenge to maintenance of immunological homeostasis in the respiratory tract. Amongst this myriad of antigens, the immune system must discriminate between innocuous components that can be tolerated by the host and potentially life‐threatening pathogens that require a rapid immune response. Dendritic cells (DC) represent the principal cell type at these sites capable of processing antigens and delivering signals that initiate tolerogenic or immunogenic immune responses. This review will discuss the role of DC at the “front‐line” of immune surveillance and homeostasis within the respiratory tract and their role in the pathogenesis of respiratory disease.
Phenotype and function of rat dendritic cell subsets
Authors: YRLID ULF , MACPHERSON GORDON
Published: September 2003

Dendritic cells (DC) comprise phenotypically‐distinct subsets that sub‐serve distinct functions in immune induction. Understanding the biology of DC subsets in vivo is crucial for the understanding of immune regulation and its perturbations in disease. This review focuses on the phenotype and functions of rat DC subsets and compares these with subsets identified in other species. Our research has concentrated on DC migrating in lymph. DC migrate constitutively from peripheral tissues to draining nodes, probably to induce/maintain tolerance to self‐ or harmless foreign antigens. After removal of mesenteric lymph nodes (MLN) in the rat, healing of afferent and efferent lymphatics permits migrating intestinal DC (iLDC) to be collected from the thoracic duct. We have shown that iLDC consist of least two subsets that differ in phenotype, in situ distribution and function. CD4+/SIRPα+ iLDC are highly immunostimulatory, but are excluded from T cell areas of MLN. In contrast, CD4−/SIRPα− iLDC are less potent stimulators of T cells, but carry material from apoptotic enterocytes to T cell areas of MLN. Similar subsets exist in both lymph nodes and spleen. It has been shown that phenotypically‐similar subsets migrate in skin‐draining lymph in cattle and sheep. We and others have shown that splenic CD4−/SIRPα− DC can phagocytose allogeneic cells in vitro, are poor stimulators of CD8+ T cells, and can lyse NK‐sensitive target cells. Although some of our data suggest that rat CD4−/SIRPα− DC may equate to murine CD8+ DC, there is at present insufficient evidence to be confident of this.
Dendritic cells in peripheral tolerance and immunity
Authors: GAD MONIKA , CLAESSON MOGENS HELWEG , PEDERSEN ANDERS ELM
Published: September 2003

Dendritic cells capable of influencing immunity exist as functionally distinct subsets, T cell‐tolerizing and T cell‐immunizing subsets. The present paper reviews how these subsets of DCs develop, differentiate and function in vivo and in vitro at the cellular and molecular level. In particular, the role of DCs in the generation of regulatory T cells is highlighted.
The role of dendritic cells in the pathogenesis of HIV‐1 infection
Authors: LORÉ KARIN , LARSSON MARIE
Published: September 2003

Dendritic cells are professional antigen‐presenting cells required for generation of adaptive immunity. These cells are one of the initial target cells for HIV‐1 infection or capture of virions at site of transmission in the mucosa. DCs carrying HIV‐1 will migrate to the lymphoid tissue where they can contribute to the dissemination of the virus to adjacent CD4+ T cells. In addition, HIV‐1–exposed DCs may have impaired antigen‐presenting capacity resulting in inadequate expansion of HIV‐1–specific T cell responses. Here, we review the infection of different subtypes of DCs by HIV‐1 and the relevance of these cells in the transmission and establishment of HIV‐1 disease. In addition, we discuss the mechanisms through which HIV‐1–DC interactions could be exploited to optimise the generation and maintenance of HIV‐1–specific T cell immunity.
Epidermal Langerhans cell migration and sensitisation to chemical allergens
Authors: CUMBERBATCH MARIE , DEARMAN REBECCA J. , GRIFFITHS CHRISTOPHER E. M. , KIMBER IAN
Published: September 2003

Epidermal Langerhans cells (LC) form part of the wider family of dendritic cells (DC; professional antigen‐processing and antigen‐presenting cells). LC are considered to serve in the skin as sentinels of the adaptive immune system, surveying the local environment and transporting foreign antigen for presentation to responsive T lymphocytes in regional lymph nodes. As such, LC play pivotal roles in the initiation of cutaneous immune responses, including immune responses to chemical allergens encountered at skin surfaces. Here we explore two aspects of LC function in the context of sensitisation to chemical allergens. The first is consideration of the cytokine and chemokine signals that regulate and counter‐regulate the mobilisation and migration of LC from the epidermis to skin‐draining lymph nodes following topical sensitisation. The second is examination of the ways in which LC may influence the polarity of induced T lymphocytes, and thereby the quality of immune responses.
Against the self: dendritic cells versus cancer
Authors: CHO HEARN JAY , BHARDWAJ NINA
Published: September 2003

The role of host defense in cancer is highly variable. Although there are cases where spontaneous cures of cancer appear to be mediated by immunologic mechanisms, malignant disease generally progresses even in patients where tumor‐specific immunity can be demonstrated. It is apparent that there are complex interactions between tumor cells and dendritic cells, the dominant antigen‐presenting cells of the immune system. Through their inhibitory actions upon dendritic cells, tumor cells can negatively regulate priming of tumor‐specific immunity. Recent work has also shown that dendritic cells have direct cytotoxic effects upon tumor cells. These interactions may impact on the efficacy of current strategies using dendritic cell‐based vaccines for tumor immunotherapy.
Clinical application of dendritic cells in cancer vaccination therapy
Authors: SVANE INGE MARIE , SOOT METTE LINE , BUUS SØREN , JOHNSEN HANS E.
Published: September 2003

During the last decade use of dendritic cells (DC) has moved from murine and in vitro studies to clinical trials as adjuvant in cancer immunotherapy. Here they function as delivery vehicles for exogenous tumor antigens, promoting an efficient antigen presentation. The development of protocols for large‐scale generation of dendritic cells for clinical applications has made possible phase I/II studies designed to analyze the toxicity, feasibility and efficacy of this approach. In clinical trials, DC‐based vaccination of patients with advanced cancer has in many cases led to immunity and in selected patients to tumor regression. However, the majority of clinical trials are still in phase I, and interpretations are hampered by pronounced variation in study design related to technical aspects of DC preparation, treatment and schedule, monitoring of immune response, and clinically relevant endpoints, including toxicity and response evaluation. This paper aims to review the technical aspects and clinical impact of vaccination trials, focusing on the generation of DC‐based vaccines, evaluation of immunologic parameters and design of clinical trials necessary to meet the need for good laboratory and clinical practice.
Calreticulin binds preferentially with B cell linear epitopes of Ro60 kD autoantigen, enhancing recognition by anti-Ro60 kD autoantibodies
Authors: STAIKOU E V , ROUTSIAS J G , MAKRI A A , TERZOGLOU A , SAKARELLOS-DAITSIOTIS M , SAKARELLOS C , PANAYOTOU G , MOUTSOPOULOS H M , TZIOUFAS A G
Published: September 2003

Calreticulin is a molecular chaperone to newly synthesized polypeptides. Previous studies suggested that calreticulin is probably a protein member of the Ro/La RNP complex. The aims of this study were (a) to investigate whether linear B cell epitopes of the Ro/La RNP complex are bound to calreticulin and (b) if the complex peptide–calreticulin is recognized specifically by anti-Ro autoantibodies. Calreticulin was isolated from either human or pig spleen using a multi-step purification method and found to interact preferentially with biotinylated peptides derived from the sequence of the Ro60 kD 175–184aa(10p) and 216–232aa(17p). The interaction of the peptide–calreticulin complex was favoured by the combination of heat treatment, divalent cations and ATP. La/SSB epitopes did not react with calreticulin. Peptides corresponding to La/SSB epitopes as well as the common epitope of Sm did not interact with calreticulin. Thirty-eight anti-Ro60 KD positive and 23 anti-Ro60 kD negative sera of patients with systemic lupus erythematosus (SLE) and primary Sjögren's syndrome (pSS) were tested. All anti-Ro60 kD positive sera bound the complex calreticulin-17p, while 95% of the same sera had activity against the complex calreticulin − 10p. Tested individually, calreticulin, pep10p and pep17p presented very low reactivity (8%, 11% and 29%, respectively) against anti-Ro60 kD positive sera. Anti-Ro60 KD negative sera did not exhibit significant reactivity either with calreticulin, 10ρ and 17ρ or with the complexes calreticulin − 10p and calreticulin-17p (<5%). These results suggest that calreticulin can induce conformation-dependent recognition of the Ro60 kD epitopes, leading eventually to their recognition by autoantibodies. This is the first time that such a relationship is shown between a chaperone protein and fragments of an intracellular autoantigen. This work also provides insights into the understanding of mechanisms for autoantibody production. Furthermore, this association can be proved useful for the development of new sensitive assays for autoantibody detection.
Childhood Helicobacter pylori infection in a murine model: maternal transmission and eradication by systemic immunization using bacterial antigen–aluminium hydroxide
Authors: MINOURA T , KATO S , OTSU S , FUJIOKA T , IINUMA K , NISHIZONO A
Published: September 2003

In humans, transmission of Helicobacter pylori is thought to occur largely during childhood. Infected mothers are generally considered to be the main source of the pathogen. However, little is known about when and how often maternal transmission of H. pylori occurs during childhood. In the present study, we examined these issues in an experimental murine model. Pregnant C57BL/6 mice, infected experimentally with H. pylori, delivered and nursed their litters. The stomachs of the infants were isolated and assessed for transmission of H. pylori. We also investigated the effect of systemic immunization using H. pylori antigen–aluminium hydroxide (AlOH) with regard to providing anti-H. pylori immunity and eradicating the maternally transmitted bacteria in infants. Polymerase chain reaction (PCR) was used to examine the presence of transmitted bacteria and their eradication. Maternal transmission of H. pylori varied widely during the nursing period, but almost all litters showed bacterial transmission at 2 weeks postpartum. Systemic immunization with bacterial antigen–AlOH eradicated the bacteria in most litters; this immunization induced a local decrease of Th2 cytokines and a local increase of Th1 cytokines in the gastric tissue, as determined by ELISA. Our results indicate that our H. pylori vaccine provides not only protection, but also eradication of the already transmitted H. pylori.
Spironolactone inhibits production of proinflammatory cytokines, including tumour necrosis factor-α and interferon-γ, and has potential in the treatment of arthritis
Authors:   THE SPIRONOLACTONE/ARTHRITIS STUDY GROUP, BENDTZEN K , HANSEN P R , RIENECK K
Published: September 2003

Evidence suggests that spironolactone, an aldosterone antagonist, has effects on many cell types independent of its binding to cytosolic mineralocorticoid receptors. We tested the effects of spironolactone on ex vivo-activated human blood leucocytes using gene expression analyses (GeneChip®, 12 000 genes) and enzyme immunoassay for quantitating secreted pro- and anti-inflammatory cytokines. Furthermore, to evaluate the safety and efficacy of spironolactone as an anti-inflammatory drug 21 patients with rheumatoid arthritis (RA), juvenile idiopathic arthritis (JIA) or other arthritides were treated for up to 22 months with 1–3 mg/kg/day. Spironolactone, at in vivo attainable doses, markedly suppressed transcription of several proinflammatory cytokines and, accordingly, inhibited release of tumour necrosis factor, lymphotoxin, interferon-γ, granulocyte-macrophage colony-stimulating factor and interleukin 6 (70–90% inhibition). Release of these cytokines was also suppressed when testing whole blood from RA patients receiving 50 mg spironolactone twice daily, indicating that pharmaceutical use of the drug may suppress the release of inflammatory cytokines. Spironolactone therapy was generally well tolerated, although treatment had to be stopped in two adults on concomitant methotrexate therapy. Sixteen patients (76%) responded favourably. American College of Rheumatology criteria (ACR)20 or better was achieved in six of nine RA patients; four reached ACR70. Eight of nine JIA patients improved. In conclusion, spironolactone inhibits production of several proinflammatory cytokines considered to be of pathogenic importance in many immunoinflammatory diseases and shows positive effect in patients with chronic arthritis. Its effect as an anti-inflammatory drug should be explored, because prolonged spironolactone therapy is reasonably safe and economically attractive compared with many modern anti-inflammatory therapies.
Homocysteine accumulates in supernatants of stimulated human peripheral blood mononuclear cells
Authors: SCHROECKSNADEL K , FRICK B , WIRLEITNER B , SCHENNACH H , FUCHS D
Published: September 2003

Moderate hyperhomocysteinaemia is associated with atherosclerosis, thrombosis and also with stroke and dementia. Elevated homocysteine concentrations are related to deficiency of folate and also vitamin-B12, as these two vitamins are essential co-factors in the remethylation of homocysteine to methionine. A causal role of homocysteine in the pathogenesis of vascular disease has been discussed over years. Immune activation appears to be involved strongly in atherogenesis as well as in other diseases found to be associated with moderate hyperhomocysteinaemia. To study a possible influence of immune stimulation on homocysteine metabolism, in vitro experiments were performed using peripheral blood mononuclear cells upon stimulation with mitogens concanavalin A, phytohaemagglutinin and pokeweed mitogen. In stimulated cells a dose-dependent increase of homocysteine concentrations was found. When cells were kept in medium supplemented with methionine, homocysteine concentrations increased further, while supplementation with folate had only a slight effect. We conclude that in supernatants of stimulated peripheral blood mononuclear cells homocysteine is accumulating. T cell activation could be involved in the development of moderate hyperhomocysteinaemia.
Childhood nephrotic syndrome in relapse is associated with down-regulation of monocyte CD14 expression and lipopolysaccharide-induced tumour necrosis factor-αproduction
Authors: CHEN S P , CHEUNG W , HENG C K , JORDAN S C , Yap H K
Published: September 2003

Interleukin-13 (IL-13) is a known modulator of monocyte function, down-regulating monocyte surface markers such as CD14 and proinflammatory cytokines. We have shown previously that lymphocyte IL-13 gene expression was up-regulated during relapses in children with steroid-responsive nephrotic syndrome (SRNS). In this study, we examined the monocyte mRNA expression and lipopolysaccharide (LPS)-stimulated intracellular production of tumour necrosis factor-α (TNF-α) and IL-8 in children with SRNS during relapse and remission. Additionally, we investigated CD14 mRNA levels, CD14 surface expression and its soluble component (sCD14) in serum. Our results showed that the percentages of TNF-α positive monocytes following LPS stimulation were significantly lower in nephrotic children in relapse (64·4 ± 13·7%) compared to remission (81·6 ± 9·0%, P < 0·005). This was associated with down-regulation of CD14 mRNA, as well as both membrane and sCD14 in patients with nephrotic relapse (82·9 ± 10·1% and 1·23 ± 0·30 µg/ml, respectively) compared to remission (93·9 ± 3·2% and 1·77 ± 0·82 µg/ml, respectively) (P < 0·003). Although we demonstrated a decrease in LPS-stimulated intracellular production of TNF-α in monocytes from patients with nephrotic relapse, we were unable to show a concomitant decrease in mRNA expression during relapses. This could be explained by down-regulation of gene expression at the translational rather than transcriptional level. In conclusion, it is conceivable that up-regulation of T-cell IL-13 production in children with active nephrotic relapse was associated with suppression of monocyte CD14 expression, down-regulating pro-inflammatory cytokine production, and could account for the increased susceptibility to bacterial sepsis seen in nephrotic children in active relapse.
Coupling of oral human or porcine insulin to the B subunit of cholera toxin (CTB) overcomes critical antigenic differences for prevention of type I diabetes
Authors: PETERSEN J S , BREGENHOLT S , APOSTOLOPOLOUS V , HOMANN D , WOLFE T , HUGHES A , DE JONGH K , WANG M , DYRBERG T , VON HERRATH M G
Published: September 2003

Our earlier investigations have demonstrated a critical difference in the efficacy of orally administered porcine compared to human or mouse insulin (no effect) in preventing type I diabetes in two distinct experimental models. Based on these findings one has to assume that certain insulins might not be suitable for the induction of oral ‘tolerance’/bystander suppression, which might be one cause for recent failures in human oral antigen trials. Here we demonstrate that coupling to the non-toxic subunit of cholera toxin (CTB) can abolish these differences in efficacy between human and porcine insulin. As expected, an added benefit was the much smaller oral antigen dose required to induce CD4+ insulin-B specific regulatory cells that bystander-suppress autoaggressive responses. Mechanistically we found that uptake or transport of insulin–CTB conjugates in the gut occurs at least partially via binding to GM-1, which would explain the enhanced clinical efficacy. Both B chains bound well to major histocompatibility complex (MHC) class II, indicating comparable immunological potential once uptake and processing has occurred. Thus, our findings delineate a pathway to overcome issues in oral antigen choice for prevention of type I diabetes.
Reconstitution of CD4+ T cell responses in HIV-1 infected individuals initiating highly active antiretroviral therapy (HAART) is associated with renewed interleukin-2 production and responsiveness
Authors: HARDY G A D , IMAMI N , SULLIVAN A K , PIRES A , BURTON C T , NELSON M R , GAZZARD B G , GOTCH F M
Published: September 2003

Reconstitution of functional CD4+ T cell responsiveness to in vitro stimuli is associated with continuous highly active antiretroviral therapy (HAART). Thirty-six antiretroviral naive patients received HAART over 16 weeks. Antigen-specific, mitogen and interleukin (IL)-2 induced lymphocyte proliferative responses and specific IL-2 and IL-4 production were assessed at each time-point, together with quantification of HIV-1 RNA load and lymphocyte populations. Reconstitution of recall responses was limited largely to persistent antigens such as Herpes simplex virus and Candida, rather than to HIV-1 or neo-antigens. Recall antigens, mitogens and IL-2-induced renewed responses were associated with in-vitro production of IL-2, but not IL-4. Differential responsiveness to low versus high concentration IL-2 stimulus increases in a stepwise manner, suggesting normalization of IL-2 receptor expression and improved functionality. These increases in in-vitro proliferative responses thus probably reflect short lived effector clones, driven by ongoing antigenic stimulus associated with persisting long-term organisms. In this context non-responsiveness to HIV-1 antigens suggests ongoing HIV-1 specific clonal T cell anergy.
cDNA cloning, expression and characterization of an allergenic L3 ribosomal protein of Aspergillus fumigatus
Authors: SAXENA S , MADAN T , MURALIDHAR K , SARMA P U
Published: September 2003

Aspergillus fumigatus (Afu) is an important fungal pathogen causing allergic and invasive respiratory disorders. A plethora of multi-functional allergens/antigens secreted by Afu have been implicated in pathogenesis. The present study was undertaken to identify and characterize novel Afu allergen/antigen by cDNA library approach. cDNA library of Afu was immunoscreened with pooled sera of allergic bronchopulmonary aspergillosis (ABPA) patients. The cDNA clone, TS1, reacting significantly with specific IgG antibodies, was selected. cDNA was subcloned and expressed in Escherichia coli. Sequencing of the cDNA revealed an open reading frame (ORF) of 1179 bases coding for a protein with an approximate molecular weight of 44 kDa. Immunoreactivity of the recombinant TS1 protein (rTS1) was evaluated by ELISA and Western blot analysis using pooled sera of ABPA patients. The rTS1 exhibited binding to specific IgG and IgE antibodies present in sera of ABPA patients. The deduced amino acid sequence showed homology to 60S ribosomal protein L3 (RpL3) of Aspergillus nidulans, Saccharomyces cerevisiae and Homo sapiens. The RpL3 of S. cerevesiae, tcm1, to which TS1 sequence shows significant homology (72% identity), is known to be responsible for conferring resistance against trichodermin (antibiotic, inhibiting protein synthesis). The present study has led to identification, cloning and expression of a 44-kDa novel allergen/antigen of Afu with sequence homology to L3 ribosomal protein with a probable role in resistance of Afu to antifungal drugs. Sixty-four per cent sequence identity of Afu RpL3 with human RpL3 and common regions in their predicted epitopes suggest a possibility of involvement of Afu RpL3 in autoimmune reactions due to molecular mimicry.
Intracellular expression ofMycobacterium tuberculosis-specific 10-kDa antigen down-regulates macrophage B7·1 expression and nitric oxide release
Authors: SINGH B , SINGH G , TRAJKOVIC V , SHARMA P
Published: September 2003

To explore the role of the 10-kDa Mycobacterium tuberculosis-specific secreted antigen (MTSA-10 or CFP-10) in modulation of macrophage function, J774 macrophages were transfected stably with DNA encoding MTSA-10. Compared to normal or mock-transfected controls, MTSA-10-expressing macrophages had markedly lower levels of co-stimulatory molecule B7·1 on their surface, while the expression of B7·2 and ICAM-1 was not affected. MTSA-transfected cells also produced significantly less microbicidal free radical nitric oxide (NO) upon stimulation with interferon (IFN)-γ, lipopolysaccharide or M. tuberculosis cell lysate. Western blot analysis revealed the absence of tyrosine-phosphorylated protein slightly larger than 112 kDa in MTSA-transfected macrophages. Moreover, the treatment of control J774 cells with protein tyrosine kinase inhibitor genistein completely mimicked the effects of transfection with MTSA-10, selectively down-regulating NO and B7·1, but not B7·2 or ICAM-1 expression. The observed MTSA-10-mediated block of B7·1 expression and NO release might contribute to the suppression of antimycobacterial response in tuberculosis.
Memory B lymphocytes determine repertoire oligoclonality early after haematopoietic stem cell transplantation
Authors: OMAZIC B , LUNDKVIST I , MATTSSON J , PERMERT J , NÄSMAN-BJÖRK I
Published: September 2003

The objective of this study was to investigate if oligoclonality of the Ig repertoire post-haematopoietic stem cell transplantation (HSCT) is restricted to memory B lymphocytes or if it is a general property among B lymphocytes. As a measure of B lymphocyte repertoire diversity, we have analysed size distribution of polymerase chain reaction (PCR) amplified Ig H complementarity determining region 3 (CDR3) in naive and memory B lymphocytes isolated from patients before HSCT and at 3, 6 and 12 months after HSCT as well as from healthy controls. We demonstrate a limited variation of the IgH CDR3 repertoire in the memory B lymphocyte population compared to the naive B cell population. This difference was significant at 3 and 6 months post-HSCT. Compared to healthy controls there is a significant restriction of the memory B lymphocyte repertoire at 3 months after HSCT, but not of the naive B lymphocyte repertoire. Twelve months after HSCT, the IgH CDR3 repertoire in both memory and naive B lymphocytes are as diverse as in healthy controls. Thus, our findings suggest a role for memory B cells in the restriction of the oligoclonal B cell repertoire observed early after HSCT, which may be of importance when considering reimmunization of transplanted patients.
Differential activation of the immune system by virulentStreptococcus pneumoniaestrains determines recovery or death of the host
Authors: MIZRACHI-NEBENZAHL Y , LIFSHITZ S , TEITELBAUM R , NOVICK S , LEVI A , BENHARROCH D , LING E , DAGAN R
Published: September 2003

Streptococcus pneumoniae infection may result in asymptomatic carriage, mucosal or invasive disease. We hypothesize that self-limiting or fatal disease outcome follows infection with S. pneumoniae differential activation of the host immune response. BALB/c and C57BL/6 mice were inoculated intranasally with S. pneumoniae serotype 3 strain WU2 and serotype 14 strain DW14 and mortality, bacterial load, pathological changes in the lungs and cytokines mRNA levels in the spleen were analysed. No differences between the C57BL/6 and the BALB/c inbred mice were observed except for the severity of their lung pathology and IL-4 expression. Infection of the two mouse strains with S. pneumoniae WU2 resulted in sepsis and death that occurred within 4 days post-inoculation. This death was preceded, in both mouse strains, in an increase over time of the lung bacterial load and bacteraemia. The lung pathology was characterized by diffuse pneumonia with marked congestion of the lungs. Analysis of mRNA expression of cytokines in the spleen revealed no alterations in tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β, interleukin (IL)-12 and interferon (IFN)-γ and induction of IL-10 and IL-4. The two strains of mice survived infection with S. pneumoniae DW14. This was accompanied by a reduction over time of lung bacterial load and bacteraemia. The lung pathology was characterized by focal lymphocyte infiltration and preserved architecture of the organ. Analysis of mRNA expression of cytokines in the spleen revealed a significant decrease in the levels of TNF-α, TGF-β, IL-12 and IFN-γ mRNA expression, which usually precedes cytokine protein expression. Interestingly, a significant increase in the levels of IL-4 mRNA expression was found in BALB/c mice only.  This study suggests that differential activation or evasion of cytokine expression by S. pneumoniae virulent strains determines disease outcome regardless of the host's immunogenetic background.
Matrix metalloproteinases production in malignant pleural effusions after talc pleurodesis
Authors: D'AGOSTINO P , RAO CAMEMI A , CARUSO R , ARCOLEO F , CASCIO A , DOLCE A , SACCO E , CANGEMI G , DI ROSA T , MOCEO P , CILLARI E
Published: September 2003

In this study we have evaluated the modifications of matrix metalloproteinases (MMPs) in malignant pleural fluids taken from patients suffering from lung cancer and treated with intrapleural talc instillation to induce pleurodesis. Furthermore, we have analysed the variations of some inflammatory mediators (C-reactive protein, α-1 antitrypsin) and of a protein (plasminogen) involved in MMP activation. In all patients the clinical improvement after talc pleurodesis was followed by a reduction in MMP-1, TIMP-1, C-reactive protein, α-1 antitrypsin and plasminogen activity. Furthermore, MMP-9 levels were variable; in fact, in some patients they were high at the beginning of treatment, in others they increased a few days after pleurodesis induction. These inhibitory effects of talc on MMP-1 and inflammatory mediators associated with the reduction of pleural effusion could constitute an effective means to evaluate the evolution of the treatment.
Methods for the in vitro determination of an individual disposition towards TH1- or TH2-reactivity by the application of appropriate stimulatory antigens
Authors: BARTH H , BERG P A , KLEIN R
Published: September 2003

In this study we performed several methods for the determination of cytokines (RT-PCR for the demonstration of cytokine mRNA and flow cytometry for the analysis of intracellular cytokines) and compared them with a recently established test system stimulating peripheral blood mononuclear cells (PBMC) with TH1- and TH2-relevant recall antigens and analysing type 1 and type 2 cytokines by ELISA. Aim of the study was therefore to evaluate the reliability of TH1/TH2 cytokine profiles in two individuals with different types of an allergic/atopic disposition: one of them showed a strong TH1/type 1-mediated tuberculin-reaction (subject A), the other (subject B) revealed elevated IgE-levels and eosinophil counts (TH2/type 2-mediated). PBMC were incubated with the type 1-antigen purified protein derivative (PPD) and the type 2-antigen tetanus-toxoid (TT) for seven days. From the comparison of ELISA with RT-PCR and flow cytometry-analysis it became evident that all three methods allowed the definition of subject A as a ‘type 1-responder’. Subject B showed a pure type 2-response in the ELISA method; PCR and flow cytometry analysis revealed the simultaneous production of type 1- and type 2-cytokines resulting in a mixed type 1/type 2-profile. Active immunization of subject A with TT at the end of the observation period of 12 months resulted in a transient shift from type 1- to a mixed type 1/type 2-profile (simultaneous PPD-induced IFN-γ- and TT-induced IL-5 production). From this pilot study based on clear cut clinical criteria concerning either a humoral or cellular immunological reactivity towards allergens/antigens it is suggested that the determination of type 1/type 2-cytokines by ELISA in supernatants of PBMC stimulated with type 1/type 2-relevant antigens is a useful approach for a better classification of ‘type1-’ or ‘type 2-responder’.
Population analysis of antiviral T cell responses using MHC class I-peptide tetramers
Authors: KOMATSU H , SIERRO S , CUERO A V , KLENERMAN P
Published: September 2003

MHC class I-peptide tetrameric complexes (‘tetramers’) have revolutionized the study of antiviral CD8+ T cell responses. They allow accurate quantification of immune responses ex vivo independent of function, with high levels of sensitivity. They have revealed unexpectedly large frequencies of ‘memory’ T cell responses against viruses such as cytomegalovirus (CMV) and Epstein–Barr virus (EBV), and provided information about their phenotypic and functional variation. However, such studies have generally concentrated on limited numbers of individuals analysed in detail. To allow larger population-based studies, we devised a method for tetramer analysis using 50–100 microlitre blood volumes in a 96-well plate format. We adapted this method to study the effect of age on responses in a cohort of nearly 600 individuals to an immunodominant HLA-A2 restricted response to CMV pp65 (NLVPMVATV). We observed the phenomenon of steady ‘memory inflation’ with age, similar to recently observed longitudinal data from murine studies. These data show that tetramers can be used as population screening tools and could be used to study age-related, geographical or seasonal effects in a number of other viral infections.
Mycophenolate mofetil inhibits differentiation, maturation and allostimulatory function of human monocyte-derived dendritic cells
Authors: ČOLIĆ M , STOJIĆ-VUKANIĆ Z , PAVLOVIĆ B , JANDRIĆ D , STEFANOSKA I
Published: September 2003

We have studied the effect of mycophenolate mofetil (MMF), a new drug used in prevention of transplant rejection, on differentiation, maturation and allostimulatory activity of human monocyte-derived dendritic cells (MDDC). MDDC were generated in vitro with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin (IL)-4 in the presence or absence of MMF. MMF reduced the number of immature MDDC in culture, dose-dependently, by inducing apoptosis and inhibited their stimulatory activity on allogeneic lymphocytes. These changes correlated with down-regulation of co-stimulatory and adhesion molecules such as CD40, CD54, CD80 and CD86. No differences were observed in mannose receptor (MR)-mediated endocytosis, measured by the uptake of fluorescein isothiocyanate (FITC)-dextran. MDDC differentiated in the presence of MMF showed significantly reduced maturation upon stimulation with lipopolysaccharide, as judged by lower expresson of CD83 and co-stimulatory molecules, lower production of tumour necrosis factor (TNF)-α, IL-10, IL-12 and IL-18 as well as lower stimulation of alloreactive T cells including naive CD4+ CD45RA+ T cells. In contrast, MDDC matured in the presence of MMF showed a more marked decrease in the FITC-dextran uptake than mature MDDC cultivated without MMF and the phenomenon correlated with down-regulation of the MR expression. These results suggest that MMF impairs differentiation, maturation and function of human MDDC in vitro, which is an additional mechanism of its immunosuppressive effect.
Expression of functional Anopheles merusα‐amylase in the baculovirus/Spodoptera frugiperda system
Authors: Effio P. C. , Folgueras‐Flatschart A. V. , Montor W. R. , Pernasetti F. M. , Pueyo M. T. , Sogayar M. C.
Published: September 2003

The Anopheles merus (Diptera, Nematocera, Culicoidea) α‐amylase gene (AmerAmy, GenBank Accession Number U01210) was amplified with its own or with the Zabrotes subfasciatusα‐amylase signal peptide (ZsAmerAmy, GenBank Accession Number AY270183) by PCR, using designed primers. The AmerAmy gene was sequenced from its promotor to the TGA codon. As a positive control, the Z. subfasciatusα‐amylase gene with its own signal peptide (ZsAmy, GenBank Accession Number AF255722) was also amplified by PCR. These three sequences were inserted into the baculovirus genome using the Bac‐to‐Bac™ system. Recombinant baculovirus preparations were used to infect Sf9 Spodoptera frugiperda insect cells. The A. merusα‐amylase was successfully expressed as an active enzyme detected mainly in cell culture supernatants.
Functional dissection of the hexamerin receptor and its ligand arylphorin in the blowfly Calliphora vicina
Authors: Hansen I. A. , Gutsmann V. , Meyer S. R. , Scheller K.
Published: September 2003

The process of receptor‐mediated uptake of hexamerin storage proteins from insect haemolymph by fat body cells is a unique feature of the class Insecta. We identified the binding domains of the hexamerin receptor and the hexamerin ligand arylphorin in the blowfly, by means of the yeast‐two‐hybrid‐system. The receptor‐binding domain of arylphorin was located within domain 3 of the arylphorin monomer. The ligand‐binding domain of the hexamerin receptor was mapped to the extreme N‐terminus of the receptor. The binding domains identified exhibit no similarity to any functional protein domains known to date. Additionally, we identified two previously unknown protein‐interactors of the hexamerin receptor. The results of this study provide further insights regarding the mechanism of the receptor‐mediated endocytosis of storage proteins in insects.
piggyBac‐mediated germline transformation in the beetle Tribolium castaneum
Authors: Lorenzen M. D. , Berghammer A. J. , Brown S. J. , Denell R. E. , Klingler M. , Beeman R. W.
Published: September 2003

The lepidopteran transposable element piggyBac can mediate germline insertions in at least four insect orders. It therefore shows promise as a broad‐spectrum transformation vector, but applications such as enhancer trapping and transposon‐tag mutagenesis are still lacking. We created, cloned, sequenced and genetically mapped a set of piggyBac insertions in the red flour beetle, Tribolium castaneum. Transpositions were precise, and specifically targeted the canonical TTAA recognition sequence. We detected several novel reporter‐expression domains, indicating that piggyBac could be used to identify enhancer regions. We also demonstrated that a primary insertion of a non‐autonomous element can be efficiently remobilized to non‐homologous chromosomes by injection of an immobile helper element into embryos harbouring the primary insertion. These developments suggest potential for more sophisticated methods of piggyBac‐mediated genome manipulation.
Haemocytes from Pseudoplusia includens express multiple α and β integrin subunits
Authors: Lavine M. D. , Strand M. R.
Published: September 2003

Cellular immune responses such as encapsulation involve the adhesion of one or more classes of haemocytes. How insect haemocytes recognize encapsulation targets as foreign or the identity of the molecules regulating haemocyte adhesion is unknown. One of the most important classes of adhesion receptors in mammalian immune cells is the integrins, which form functional heterodimers through different combinations of α and β subunits. Prior studies with the moth Pseudoplusia includens indicated that encapsulation depends on two classes of haemocytes called granulocytes and plasmatocytes. Here we report the cloning and identification of three α integrin subunits (αPi1–3) and one β subunit (βPi1) from P. includens. Northern blot analysis indicated that all four subunits are expressed in granulocytes and that three of the four subunits are expressed in plasmatocytes. Quantification of transcription patterns using real‐time PCR revealed that expression of αPi2 and βPi1 increased in granulocytes and plasmatocytes when binding to a foreign surface or forming a capsule. αPi2 transcription in plasmatocytes was further increased by granulocyte conditioned medium, plasmatocyte spreading peptide, and the integrin recognition peptide RGD. Collectively, these results suggest that one or more integrins play an important role in regulating haemocyte adhesion during encapsulation.
Fall armyworm FR sequences map to sex chromosomes and their distribution in the wild indicate limitations in interstrain mating
Authors: Nagoshi R. N. , Meagher R.
Published: September 2003

The fall armyworm, Spodoptera frugiperda, consists of two host strains (rice and corn) that differ in developmental, physiological and behavioural characteristics. However, because the strains are morphologically indistinguishable the investigation of strain‐specific behaviour, particularly in the wild, is very difficult. This has spurred the isolation of diagnostic molecular markers. FR sequences are tandem‐repeat genetic elements found in large clusters only in the rice strain. To facilitate their use as a strain marker we genetically mapped FR clusters and found they localized to the sex chromosomes. This represents one of the first examples of chromosome mapping in fall armyworm. The FR sequence and a strain‐specific mitochondrial marker were then used to examine the distribution of different marker combinations in field specimens. These studies identified significant barriers to interstrain mating in the wild, specifically that corn strain females rarely, if ever, mate with rice strain males. The data also suggest that only a genetically distinct subset of the overwintering rice strain population in Florida annually migrate to Georgia. These studies demonstrate that the availability of genetically characterized molecular markers for strain identity makes possible studies on fall armyworm biology in the wild previously considered unfeasible.
Developmental expression and gene/enzyme identifications in the alpha esterase gene cluster of Drosophila melanogaster
Authors: Campbell P. M. , De Q. Robin G. C. , Court L. N. , Dorrian S. J. , Russell R. J. , Oakeshott J. G.
Published: September 2003

Here we show how the 10 genes of the alpha esterase cluster of Drosophila melanogaster have diverged substantially in their expression profiles. Together with previously described sequence divergence this suggests substantial functional diversification. By peptide mass fingerprinting and in vitro gene expression we have also shown that two of the genes encode the isozymes EST9 (formerly ESTC) and EST23. EST9 is the major ‘alpha staining’ esterase in zymograms of gut tissues in feeding stages while orthologues of EST23 confer resistance to organophosphorus insecticides in other higher Diptera. The results for EST9 and EST23 concur with previous suggestions that the products of the alpha esterase cluster function in digestion and detoxification of xenobiotic esters. However, many of the other genes in the cluster show developmental or tissue‐specific expression that seems inconsistent with such roles. Furthermore, there is generally poor correspondence between the mRNA expression patterns of the remaining eight genes and isozymes previously characterized by standard techniques of electrophoresis and staining, suggesting that the alpha cluster might only account for a small minority of the esterase isozyme profile.