Vol. 49, No. 1, 2001

DNA Tumor Vaccines

ANTHONY P. WLAZLO and HILDEGUND C. J. ERTL (The Wistar Institute, 3601 Spruce Street, Philadelphia, PA 19104, USA)

Abstract. A new generation of vaccines are being developed to induce immune responses that fight off infectious agents, or erradicate cancerous cells. The new vaccines are based on a plasmid vector, which in transfected mammalian cells cause constitutive high-level expression of the target antigen. Expression of the target antigen, in turn, can induce a full-range of immunologic responses, including cell-mediated killing, cell-mediated cytokine release and the production of antigen-specific antibodies. Through molecular techniques, these nucleic acid vaccines can be enhanced to increase target antigen expression and faciliatate antigen presentation. Additionally, genetic adjuvants expressed simultaneously with the target antigens can induce the immune responses to disease-associated antigens. The ease with which these genetic vaccines can be generated and the potency of their ability to generate immune-mediated responses make them highly effective, which creates hope for developing effective treatment and prevention of various diseases, most notably cancer.

Keywords: DNA vaccine; T cells; cancer; immunity; cytokines

49z101

CD8+ T cell Suppressor Factors and the Control of Infection, Replication and Transcription of Human Immunodeficiency Virus

KAREN F. T. COPELAND (Ottawa Hospital Research Institute, Ottawa, Ontario, Canada)

Abstract. CD8+ T cells have been shown to produce factors which modulate HIV-1 replication in both T cells and monocytic cells. Examination of the literature reveals that this modulation may occur by the production of -chemokines which block viral entry. However, another CD8+ T cell-derived activity targets the replication of HIV-1 at the level of transcription. CD8+ T cell factors strongly suppress replication and transcription in T cells and T cell lines, and in contrast, the factors enhance both replication and transcription in cells of the monocyte/macrophage lineage. The enhancement of transcription and replication by CD8+ T cell factors is induced by increased production of TNF by the macrophages. The enhancement is sensitive to pertussis toxin, indicating a G protein-coupled pathway. Thus, CD8+ T cells produce factors which mediate effects on transcription and replication of HIV-1 in a cell type-dependent manner. In this review a summary of the effects of chemokines and CD8-derived factors on HIV-1 transcription and replication is presented. The virus-host cell interactions that participate in the persistent replication of HIV in macrophages and the suppression of these functions in T cells require definition.The identification of CD8+ T cell factors which exert these controls on HIV-1 may lead to promising new therapies for HIV infection.

Keywords: CD8+ T cell; transcription; replication; HIV-1; macrophage; pertussis toxin.

49z102

Phage Display as a Tool for Rapid Cloning of Allergenic Proteins

ULRICH APPENZELLER, KURT BLASER and RETO CRAMERI

Abstract. Allergic diseases represent an immune disorder associated with the production of immunoglobulin E (IgE) against normally innocuous antigens (allergens). Almost 20% of the population in industrialised countries suffer from Type I allergic symptoms like allergic rhinitis, conjunctivitis, urticaria or asthma. Although the mechanisms responsible for these allergic reactions are quite well understood, knowledge about the repertoire of molecules able to elicit Type I symptoms is still limited. To clone and characterise entire allergen repertoires from complex allergenic sources in a fast and efficient way, new technologies are required. The phage surface display of cDNA libraries described here has proven to be a versatile cloning system to selectively isolate allergens physically linked to their genetic information. The screening of cDNA libraries displayed on phage surface with immobilised serum IgE from allergic patients reduces the time required for the selection of candidate clones to a few weeks. Robot assisted high throughput screening of enriched library provides a fast and cost-effective way to isolate complete allergen repertoires. The biotechnological production of recombinant allergens derived from these sequences bear a high potential for the improvement of the diagnosis of allergic diseases.

Keywords: cloning; cDNA; phage surface display; recombinant allergens; allergy.

49z103

Inhibition of Angiogenesis in the Treatment of Tumors

KAZIMIERZ OSTROWSKI and AGNIESZKA KINSNER

Abstract. Angiogenesis is essential for tumor progression, growth and metastases. Many substances present in a normal organism can inhibit or stimulate the process of new vessel formation in tumors. The use of natural or synthetic angiogenesis inhibitors as anticancer drugs is currently under intense investigation. Such agents can have lower toxicity and are less likely to generate drug resistance than conventional cytotoxic drugs. Clinical trials are now underway to develop optimum treatment strategies for antiangiogenic drugs. This paper reviews the present achievements in preclinical and clinical studies with antitumor drugs based on inhibitors of angiogenesis.

Keywords: inhibitors; angiogenesis; cancer treatment.

49z104

Immunomodulatory Role of the Corticotropin Releasing Factor

MARKO RADULOVIC and JOACHIM SPIESS

Abstract. Corticotropin-releasing factor (CRF) was originally identified as a hypothalamic peptide which stimulates secretion of the hypophyseal adrenocorticotropic hormone. CRF exhibits its actions through G protein-dependent seven-membrane-domain receptors. Two subtypes of CRF receptors (CRFR1 and CRFR2) have been characterized thus far. CRF and its receptors were found in a number of brain regions, where they function by neuromodulation, and also in several peripheral organs. Besides CRF, another naturally occurring CRF-like peptide, urocortin, has been characterized. In the immune system, CRF and CRFR1 have so far been detected at both mRNA and protein lelvels in several lymphoid organs and at sites of inflammation. Locally injected CRF was shown to modulate the severity of inflammation. This effect was not only a result of hemodynamic changes known to be induced by CRF or by activation of the hypothalamo-pituitary adrenal axis, as CRF-binding sites were also found on immune cells. CRF was shown to directly modulate secretion of cytokines and neuropeptides, proliferation, chemotaxis and degranulation of purified macrophage and lymphocyte populations in vitro . The presence of functional CRFR was more recently demonstrated also on polymorphonuclear cells and significant amounts of CRF were shown to be produced in lymphoid organs, or delivered to lymphoid organs by peripheral nerves. Taken together, the experimental results obtained so far strongly point to the importance of CRF as a signaling molecule in lymphoid tissues and at the sites of inflammation.

Keywords: corticotropin-releasing factor; inflammation; neutrophil; stress; immune system.

49z105

Expression and Functional Significance of CTLA-4, a Negative Regulator of T Cell Activation

AGATA KOSMACZEWSKA, LIDIA CISZAK, DOROTA BOĆKO and IRENA FRYDECKA

Abstract. The generation of an effective immune response involves antigen-specific T cell expansion and differentiation of effector function. T cell activation requires at least two distinct signals, including signaling via the Ag-specific TCR and a costimulatory pathway. Antigen stimulation of T cells can lead either to a productive immune response characterized by proliferation, differentiation, clonal expansion and effector function or, in absence of appropriate costimulation, to a state of long-lasting unresponsiveness termed anergy. Anergic T cells fail to proliferate and secrete cytokines in response to secondary stimulation. The interaction between costimulatory molecule CD28 on T cells with members of the B7 family on APC results in upregulation of T cell proliferation, cytokine production and induces the expression of the anti-apoptotic protein Bcl-xl. Based of those findings, the two-signal requirement model for T cell activation is today generally accepted. The negative regulatory mechanisms during T cell activation are not well understood, but they are crucial for the maintainance of lymphocyte homeostasis. For several years the functional role of the enigmatic CD28 homologue CTLA-4 (cytotoxic T lymphocyte antigen-4) in T cell activation has been both obscure and conteroversional. CTLA-4 was initially proposed to provide a costimulatory signal in conjunction with TCR/CD3 signaling. Today we know that CD28 and CTLA-4 molecules may have diametrically opposed functions: signaling via CD28, in conjunctive with TCR, is required for T cell activation, while signaling via CTLA-4 is a negative signal that inhibits T cell proliferation. How the T cell integrates signals through the TCR/CD3 complex, CD28 and CTLA-4 to initiate, maintain and terminate antigen-specific immune response is actually not fully clarified. In this review, we will focus on the emerging role of CTLA-4 as a negative regulator of T lymphocyte activation and its role in dynamic interplay of activatory and inhibitory signals.

Keywords: CTLA-4; T cell activation; negative regulation.

49z106

The Murine Ly49 Family: Form and Function

ANDREW P. MAKRIGIANNIS and STEPHEN K. ANDERSON

Abstract. The activity of natural killer (NK) cells is regulated by surface receptors that recognize class I MHC. Murine NK cells express a large family of lectin-related receptors (Ly49s) to perform this function, while human NK cells utilize a separate group of proteins containing Ig-related domains (KIRs). Although these receptor families not structurally related, the Ly49 family appears to be the functional equivalent of human KIRs, since it uses similar signal transduction pathways for either activation or inhibition of NK cell function. Therefore, lessons learned from the study of the murine MHC class I receptor system may be relevant to human NK function. This review summarizes the current state of knowledge of the Ly49 family.

Keywords: NK cells; MHC class I; receptors; signal transduction; Ly49; gene family.

49z107

A Method for Directly Determining the Number of Dendritic Cells and for Evaluation of Their Function in Small Amounts of Human Peripheral Blood

Sergiusz Markowicz, Henryk M. Skurzak and Jan Walewski

Abstract. Bone marrow-derived dendritic cells (DC) are highly potent antigen-presenting cells capable of initiating primary responses of naive T lymphocytes to antigen. Studies on DC in disease have been impeded by the lack of a defined method for accurate DC counting and for evaluation of their function in a small amount of blood. In order to detect and enumerate DC in whole peripheral blood preparations, we applied a direct two-color immunofluorescence method. Blood from healthy donors was stained with a mixture of FITC-conjugated monoclonal antibodies (moAbs) recognizing lineage-associated molecules (CD3, CD14, CD16, CD20, CD57) and PE-conjugated anti-HLA-DR moAb. DC were identified as lineage marker negative (lin-), HLA-DR highly positive cells. The mean percentage of these cells present in peripheral blood leucocytes (PBL) was 0.54%, and the mean absolute DC count was 31.4 x 106/l of blood. DC stained directly in whole blood were heterogeneous with regard to their expression of CD2 and CD4 molecules, and did not express CD80 and CD83 molecules. Expression of CD80 and CD83 on DC was induced following a multistep isolation procedure, including overnight culture. We demonstrated a significant primary proliferative response to keyhole limpet hemocyanin (KLH) in cultures of peripheral blood mononuclear cells (PBMNC). Since primary proliferative response to neoantigens is entirely dependent on DC as antigen-presenting cells, the cultures of unseparated PBMNC stimulated with KLH can be used to evaluate DC function in a relatively simple test. This test does not require previous isolation of DC and T lymphocytes and, therefore, can be performed on a small amount of blood. The elaborated flow cytometric method of DC counting in blood and the proliferative test of DC-dependent primary response to neoantigen are currently being applied in an ongoing study on the effect of chemotherapy on DC number and function in cancer patients.

Keywords: dendritic cells; dendritic cell phenotyping; primary proliferative response.

49z108

Malformations of Angiogenesis in the Low Differentiated Human Carcinomas. Immunohistochemical Study

ALBINA ŻÓŁTOWSKA, JAN STĘPIŃSKI, BARBARA LEWKO, BARBARA ZAMORSKA, ANDRZEJ ROSZKIEWICZ, KRYSTYNA SERKIES and WIESŁAW J. KRUSZEWSKI

Abstract. Our previous observations showed that the perivascular mesenchyma of the thin-walled vessels (capillaries) in cancers may be the source of organ-specific stem cells. We suggested that the cells forming vascular channels in altered stroma participate in the tumor development. This study was designed to examine the distribution of the vessels and their appearance in the breast, lung and colon cancers. Using immunohistochemical methods, we have shown that in the low differentiated tumors both CD31 and factor VIII antigens may be expressed in capillaries chiefly on the periphery of neoplastic foci. Many of these vessels were discontinuous, with interruptions or unformed tubules. Sporadically, CD31 protein and factor VIII antigens were not expressed in capillaries inside the very low differentiated cancer cases. It is difficult to assess by immunohistochemichal means whether the vascular malformations are the primary or secondary phenomena in the malignancy and why these abnormalities were especially visible in some low differentiated cancers.

Keywords: cancer; malformation of angiogenesis; immunohistochemistry.

49z109

TNF-a, IL-6 and Their Soluble Receptor Serum Levels and Secretion by Neutrophils from Cancer Patients

EWA JABŁOŃSKA (Department of Immunopathology), MAREK KILUK, WŁODZIMIERZ MARKIEWICZ (Department of Surgery, Regional Center of Oncology), LESZEK PIOTROWSKI, ZYTA GRABOWSKA (Department of Oral and Maxillofacial Surgery) and JAKUB JABŁOŃSKI (Department of Toxicology, Medical Academy of Bialystok, Kilińskiego 1, 15-230 Białystok, Poland)EWA JABŁOŃSKA (Department of Immunopathology), MAREK KILUK, WŁODZIMIERZ MARKIEWICZ (Department of Surgery, Regional Center of Oncology), LESZEK PIOTROWSKI, ZYTA GRABOWSKA (Department of Oral and Maxillofacial Surgery) and JAKUB JABŁOŃSKI (Department of Toxicology, Medical Academy of Bialystok, Kilińskiego 1, 15-230 Białystok, Poland)

Abstract. Simultaneous evaluation of cytokines and their soluble receptor production and serum levels can be helpful in understanding the local and systemic immune response of a tumor-bearing host. In the present study we examined the serum levels of TNF-(, IL-6 and their soluble receptors sTNFRp55, sTNFRp75 and sIL-6R compareded with their production by the polymorphonuclear neutrophils (PMN) from cancer patients. Examinations were carried out in patients with adenocarcinoma breast cancer and squamous cell carcinoma of the oral cavity and related to the clinical course and to different phases of therapy. Secretion of IL-6, sTNFRp55 and sTNFRp75 by PMN appeared to be dependent on tumor type, clinical progression of disease as well as on therapy, suggesting a significant role of these cells at different phases of the immune response to cancer associated with these mediators. Changes in values of TNF-(, IL-6 and their soluble receptors in sera of both cancer groups, dependent on tumor type, clinical progression and cancer therapy, could have a diagnostic and prognostic role in cancer disease.

Keywords: tumor necrosis factor-a (TNF-a); interleukin 6 (IL-6); soluble IL-6 receptor; soluble TNF receptor; breast cancer; oral cavity cancer.

49z110

Cytokine Production in Whole Blood Cell Cultures of Patients with B-Lineage Acute Lymphoblastic Leukemia. The Influence of Granulocyte-Macrophage Colony Stimulating Factor

TERESA KAMIŃSKA, MARTYNA KANDEFER-SZERSZEŃ, IWONA HUS, ANNA DMOSZYŃSKA

Abstract. We investigated the levels of 6 different cytokines in the sera of 10 newly diagnosed patients with B-cell lineage acute lymphoblastic leukemia (ALL) and detected a significant increase in IL-6 and IFN-( serum levels in comparison to that of healthy controls. Whole blood cell cultures of 10 ALL patients and 20 control individuals were induced with classical cytokine inducers, such as virus, PHA and LPS, and their ability to produce 9 different cytokines was compared. Blood cells of ALL patients produced significantly less IL-1(, IL-1(, IL-10 and TNF-( than control cells and not significanly lower levels of IL-6, but comparable with control levels of IL-2, IL-4. rHuGM-CSF added to cell cultures 24 hr before induction significantly enhanced the production of IL-1(, IL-1( and TNF-( in controls, but only IL-1( and IL-1( in the blood cell cultures of patients with ALL. GM-CSF did not significantly influence the production of IFN-(, IFN-(, IL-2, IL-4 and IL-10 in the control cells and the cells of ALL patients. The patients examined differed not only in the expression of CD10 and CD34 antigens on blast cells, but also in the reaction to GM-CSF treatment, which was found as very high standard deviation values. We suppose that these differences can partially explain the different effects of GM-CSF when used to ameliorate neutropenia of ALL patients after chemotherapy and to reduce the incidence of microbial infections.

Keywords: GM-CSF; acute lymphoblastic leukemia; IL-1; IL-2; IL-4; IL-6; IL-10; TNF-a; IFN-a; IFN-a.

Vol. 49, No. 2, 2001

Review

Extrathymic Pathways of T Cell Differentiation
TORU ABO

Abstract. Department of Immunology, Niigata University School of Medicine, Niigata, Japan

Keywords: It is known that the liver is a major hematopoietic organ at fetal stages, but the hematopoiesis of this organ ceases of birth. However, the liver is still found to comprise c-kit+ stem cells and gives rise to extrathymic T cells, NK cells, and even granulocytes after birth. Extrathymic T cells generated in the liver of mice are identified as intermediate TCR (TCRint) cells, which include the NK1.1+TCRint (i.e. NKT cells) and NK1.1-TCRint subsets. Although extrathymic T cells are few in number during youth, they increase in number with advancing age. The number and function of extrathymic T cells are also elevated under conditions of stress, infections, malignancy, pregnancy, autoimmune disease, chronic GVH diseases, etc. Under these conditions, the mainstream of T cell differentiation in the thymus, which produces conventional T cells, is inversely suppressed. Extrathymic T cells comprise self-reactive forbidden clones and mediate cytotoxicity against abnormal self-cells. Therefore, they might be beneficial for the elimination of such cells. However, over-activation of extrathymic T cells might be responsible for the onset of certain autoimmune disease.

49z201
The Receptors Regulating Natural Cytotoxic Effector Functions
MARIA NIKOLOVA (National Center of Infectious and Parasisitic Diseases, 1504 Sofia, Bulgaria), LAURENCE BOUMSELL and ARMAND BENSUSSAN (INSERM 448, 94000 Creteil, France)

Abstract. Natural cytotoxic effector functions are regulated by a multitude of opposing signals provided by immunoglobulin and lectin-like functional molecules. While inhibitory receptors possess ITIM cytoplasmic sequences recruiting tyrosine phosphatases, activatory receptors require association with accessory ITAM-bearing molecules. One considerable group of natural cytotoxic cell receptors are specific for classical and non-classical class I antigens and detect both qualitative and quantitative changes in the autologous MHC-I phenotype. Non-MHC-I specific receptors provide signaling in the absence of MHC-I antigens or in response to not well-known stress-induced antigens. NK cell receptors may equally participate in the regulation of target cell functions through contact or soluble mediator-dependent mechanisms. The identification of NK cell-regulating molecules has lead to the elucidation of more general principles underlying immune homeostasis.

Keywords: natural cytotoxicity; NK cells; KIR; MHC-I; BY55.

49z202
Neuropeptides as Modulators of Macrophage Functions. Regulation of Cytokine Production and Antigen Presentation by VIP and PACAP
DOINA GANEA (Department of Biological Sciences, Rutgers University, Newark, NJ 07102, USA) and MARIO DELGADO (Departamento Biologia Celular, Facultad de Biologia, Universidad Complutense, Madrid 28040, Spain)

Abstract. VIP, and the structurally related neuropeptide PACAP, present in the microenvironment of lymphoid organs, modulate the function of inflammatory cells through specific receptors. VIP and PACAP inhibit the production of the pro-inflammatory agents and stimulate the production of anti-inflammatory cytokines in activated macrophages. The effect is mediated through specific receptors, and involves shedding of the CD14 LPS receptor and the transcriptional regulation of cytokine genes through effects on de novo expression or nuclear translocation of NFkB, CREB, c-Jun, and IRF-1. The in vivo administration of VIP/PACAP results in a similar pattern of cytokine modulation, which presumably mediates the protective effect of VIP/PACAP in a high-endotoxic murine model for septic shock. VIP/PACAP reduce the expression of the costimulatory B7.1/B7.2 molecules, and the subsequent stimulatory activity for T helper cells in stimulated macrophages. In contrast, in unstimulated macrophages, VIP/PACAP induce specific B7.2 expression and promote Th2 cell differentiation. We propose that VIP/PACAP act as endogenous factors that regulate immune homeostasis, and that the physiological consequences of the VIP/PACAP presence in the immune microenvironment depends on the timing of the neuropeptide release and the activation stage of the neighboring immune cells.

Keywords: neuropeptides, macrophages, cytokines, B7 molecules, Th1/Th2 cells, VIP/PACAP.

49z203
Modeling the Meta-Dynamics of Lymphocyte Repertoires
RAMIT MEHR (Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel)

Abstract. The complexity of biological systems, and the explosion of the quantity of biological information which is rapidly becoming available from experimental and clinical studies, necessitate the use of theoretical tools, namely, mathematical and computational modeling. The vertebrate adaptive immune system, with its learning and memory capabilities, is a particularly rich source of modeling challenges. Most difficult within this area is the study of lymphocyte repertoires – the generation of their diversity and the forces that shape the ever-changing dynamics of lymphocyte clones. I review several examples of problems in lymphocyte repertoire modeling, demonstrate the types of solutions employed, and highlight the contribution of these theoretical studies to immunological research.

Keywords: lymphocyte repertoires; lymphocyte development; mathematical model; computer simulation.

49z204
The Molecular Mechanisms of Post-Adhesive Transmigration of T Cells
JUN-ICHI MASUYAMA (Division of Rheumatology and Clinical Immunology, Jichi Medical School, Minamikawachi-machi Tochigi, 329-0498, Japan)

Abstract. Leukocyte extravasation is an essential phenomenon in inflammatory responses of the body. However, less is known about the mechanisms of transendothelial migration of leukocytes subsequent to their adhesion to the endothelium. It could be considered that at least three different cellular responses participate in the transmigration of adherent leukocytes: 1) polarization of adherent cells in cell shape, 2) interactions between adherent cells and molecules bound to the endothelial surface to stimulate migration through the junction between adjacent endothelial cells, and 3) co-ordination with endothelial cells to open the junction. Molecules involved in these events are discussed in this review.

Keywords: transmigration; endothelial cells; T cells; molecular mechanism.

49z205
Molecular Mechanisms of CD40 Signaling
GAIL A. BISHOP and BRUCE S. HOSTAGER (Department of Microbiology, The University of Iowa, Iowa City, IA 52240, USA)

Abstract. CD40, a member of the growing tumor necrosis factor receptor (TNF-R) family of molecules, functions as a transmembrane signal receptor in both hematopoietic and non-hematopoietic cell types, although its physiological roles are less well understood in the latter. Much has been learned over the past decade about the role of CD40 signaling in various cellular functions. In addition, some of the molecular events which occur subsequent to CD40 engagement have been characterized, although much remains to be understood. This review will summarize the known important biological roles of CD40, and discuss what is currently known about how CD40 signals.

Keywords: CD40; lymphocyte activation; sugnal transduction; B cell; tumor necrosis family; recptor family.

49z206

Clinical Immunology

Alcohol-Related Cirrhosis with Pancreatitis. The Role of Oxidative Stress in the Progression of the Disease
AGNIESZKA SZUSTER-CIESIELSKA (Department of Virology and Immunology, Maria Curie-Skłodowska University, Akademicka 19, 20-033 Lublin, Poland), JADWIGA DANILUK (Clinic and Department of Gastroenterology, University Medical School, Jaczewskiego 8, 20-950 Lublin, Poland) and MARTYNA KANDEFER-SZERSZEŃ (Department of Virology and Immunology, Maria Curie-Skłodowska University, Akademicka 19, 20-033 Lublin, Poland)

Abstract. To assess the level of oxidative stress, measured as prooxidant-antioxidant imbalance in the blood of patients with alcohol-related injury of the liver and pancreas, we determined superoxide ion (O2•-) production by neutrophils isolated from the peripheral blood of 3 groups of patients. Patients with compensated alcoholic liver cirrhosis (n=16), with alcoholic chronic pancreatitis (n=20), and with concomitant cirrhosis and pancreatitis (n=10) were included in this study. All patients had consumed at least 70 g of pure alcohol per day over 5 years. They had not abstained before admission to hospital. The control group consisted of 16 healthy non-alcohol-abusive subjects. As antioxidative enzymes (AOE) present in sera play a very important role in the regulation of plasma ROS levels and in the protection of plasma compounds against ROS action, we also examined serum activity of CAT, SOD (total activity) and GPx serum concentration. Neutrophils of patients with concomitant alcoholic liver cirrhosis and pancreatitis exhibited, similarly to the neutrophils of patients with chronic alcoholic pancreatitis, an enhanced ability to produce superoxide anions in vitro. In contrast, neutrophils of patients with alcoholic liver cirrhosis exhibited a defect in resting and PMA-induced superoxide anion production. The AOE activity in the sera of patients was also significantly changed. Total SOD activity was enhanced in all groups of patients with alcoholic liver cirrhosis, chronic pancreatitis and with concomitant injury of both organs. CAT activity was only increased in the sera of patients with liver cirrhosis or pancreatitis, but not in the patients with concomitant cirrhosis and pancreatitis. GPx concentration was only diminished in the patients with chronic pancreatitis. It seems likely that oxidative stress, defined as the imbalance between prooxidant and antioxidant activity, is highest in the blood of patients with chronic pancreatitis and, especially, in patients with concomitant liver cirrhosis and pancreatitis.

Keywords: alcohol; cirrhosis; pancreatitis; superoxide anion; catalase; glutathione peroxidase; superoxide dismutase.

49z207
Lactoferrin Regulates Proliferative Response of Human Peripheral Blood Mononuclear Cells to Phytohemaggultinin and Mixed Lymphocyte Reaction
MICHAŁ ZIMECKI, DARIUSZ STĘPNIAK, AGNIESZKA SZYNOL (Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Weigla 12, 53-114 Wrocław, Poland) and MARIAN L. KRUZEL (Department of Integrative Biology, Pharmacology and Physiology, Health Science Center, Medical School, University of Texas, PO BOX 20708, Houston TX 77225, USA)

Abstract. The aim of this study was to investigate the effects of bovine lactoferrin on the proliferative response of human blood lymphocytes induced by PHA and alloantigens in two-way mixed lymphocyte culture at a broad range of BLF concentrations (1.5 – 50 ?g/ml). We found that the effects of BLF in both experimental models were differential and depended on an individual reactivity of lymphocytes with respect to mitogen or alloantigen and BLF concentration. Generally, lymphocytes from donors reactive to BLF exhibited higher proliferation indices compared to nonreactive individuals. Low BLF doses were regulatory and higher ones mostly inhibitory. MLR was in most cases inhibited by all doses of BLF, some MLR stimulated and other not affected by BLF. We conclude that basis for the differential action of BLF is its ability to sense the activation status of lymphocytes. The resultant effects of BLF are probably mediated by monocytes and cytokines.

Keywords: bovine lactoferrin; PBMC; proliferation; PHA; MLR.
Proinflammatory Cytokine Inhibitors, TNF-a and Oxidative Burst of Polymorphonuclear Leukocytes in Pathogenesis of Sepsis in Newborns
JANUSZ PIOTR SIKORA, DANUTA CHLEBNA-SOKÓŁ (Department of Pediatric Propedeutics, Institute of Pediatrics, Medical University of Łódź), IWONA DĄBROWSKA, DARIUSZ LIPCZYŃSKI (Department of Intensive Care and Anesthesiology, 4th Pediatric Clinical Hospital, Medical University of Łódź) and SŁAWOMIR CHRUL (Department of Child Diseases, Institute of Pediatrics, Medical University of Łodź, Sporna 36/50, 91-738 Łódź, Poland)

Abstract. This study was to evaluate the levels of the proinflammatory cytokine tumor necrosis factor alpha (TNF-a) and the cytokine inhibitors soluble TNF-a receptor (sTNFR) and IL-1 receptor antagonist (IL-1 ra) as well as the intensity of oxidative metabolism of peripheral blood polymorphonuclear leukocytes in the course of sepsis in newborns. An increase of TNF-a, sTNFR and IL-1 ra concentrations was found in the blood serum of the patients at the time of diagnosis. This was further accompanied by polymorphonuclear leukocyte stimulation and, as a consequence of prolonged bacterial antigen stimulation, functional exhaustion of these cells and their diminished oxidative metabolism was observed. Within the same time period, an enhanced expression of p55 and p75 TNF-a receptors on polymorphonuclear leukocyte cell surfaces was found. It was indicated that the applied pharmacotherapy caused a decrease of the initially elevated concentrations of TNF-a and proinflammatory cytokine inhibitors (sTNFR, IL-1 ra). The intensive therapy of sepsis was associated with the increased oxidative burst of polymorphonuclear leukocytes along with the decrease of p55 and p75 expression on their cell surfaces.

Keywords: sepsis; TNF-a; cytokine inhibitors; oxidative burst; neutrophils.

49z209

Immunochemistry

Structure and Serological Characterization of an Ne-[(R)-1-carboxyethyl]-L-lysine-Containing the O-Chain of the Lipopolysaccharide of Proteus mirabilis O13
ANNA S. ŚWIERZKO, MACIEJ CEDZYŃSKI, ANDRZEJ ZIÓŁKOWSKI, WIESŁAW KACA (Centre of Microbiology and Virology, Polish Academy of Sciences, Lodowa 106, 93-232, Łódź, Poland), SOF’YA N. SENCHENKOVA, ANDREI V. PEREPELOV, YURIY A. KNIREL (N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, Russia)

Abstract. In this paper we present the structure and describe serological properties of the O-specific polysaccharide of Proteus mirabilis O13 lipopolysaccharide, which contains a unique component, an amide of D-galacturonic acid (D?GalA) with an unusual amino acid Ne-[(R)-1-carboxyethyl]-L-lysine (alaninolysine, AlaLys). Selective chemical degradations of either GalA or AlaLys resulted in the loss of the serological reactivity of the polysaccharide with anti-O serum against P. mirabilis O13. Neither synthetic stereoisomers of AlaLys nor the isolated amide of GalA with AlaLys inhibited the reaction of the O-antiserum with the homologous lipopolysaccharide. The O-antiserum did not cross-react with the lipopolysaccharide of Providencia alcalifaciens O23 containing an amide of D-glucuronic acid with AlaLys. These data showed that both uronic acid and amino acid components of the amide play an important role in manifesting the P. mirabilis O13-specificity, but the full specific epitope also includes also another OPS component(s). A cross-reactivity of anti-O13 serum with some other P. mirabilis strains was observed and attributed to a common heat-stable antigen(s) different from the lipopolysaccharide

Keywords: Proteus mirabilis; lipopolysaccharide; O-specific polysaccharide; serological reactivity; Ne-[(R)-1-carboxyethyl]-L-lysine

49z210
Neural Cell Adhesion Molecule in Breast, Colon and Lung Carcinomas
ALBINA ŻÓŁTOWSKA, JAN STĘPIŃSKI, BARBARA LEWKO (Department of Immunopathology), KRYSTYNA SERKIES (Department of Oncology and Radiotherapy), BARBARA ZAMORSKA (Department of Immunopathology), ANDRZEJ ROSZKIEWICZ, EWA IŻYCKA-ŚWIESZEWSKA (Department of Pathomorphology) and WIESŁAW J. KRUSZEWSKI (Department of Surgical Oncology Medical University Gdańsk, Dębinki 7, 80-211 Gdańsk, Poland)

Abstract. Neural cell adhesion molecules (NCAM) play an important role in embryogenesis and in some tumors, especially of neuroectodermal origin. In this study, eighteen cases of invasive breast carcinoma, seven cases of sigmoid colon carcinomas and seventeen cases of the non-small cell lung carcinoma were immunostained for NCAM. The NCAM expression, usually focal, was observed in some cases only. NCAM was expressed in the membranes, in a fine granular pattern. In three cases of breast cancers, also cytoplasmic localisation of NCAM was observed, which may suggest its cytoplasmic formation. Furthermore, in three cases expression of NCAM in histologically normal ductal lobular units adjacent to invasive breast cancers without presence of this antigen in cancer tissue was observed. The immunostaining was weak or absent in sigmoid colon carcinomas. In this study we confirm observation of some authors that NCAM expression occurs in some cases of non-small cell lung carcinomas.

Keywords: CD56 (NCAM); breast ca; sigmoid colon ca; squamous cell lung ca.

49z211

Vol. 49, No. 3, 2001

Review

Lipoxins and Aspirin-Triggered 15-epi-Lipoxins are Endogenous Components of Antiinflammation: Emergence of the Counterregulatory Side
CHARLES N. SERHAN (Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesiology, Perioperative and Pain Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA)

Abstract. Eicosanoids are known to play important roles in cell-cell communications and as intracellular signals that are critical components of multi-cellular responses such as acute inflammation and reperfusion injury. Recent findings have given rise to several new concepts that are reviewed here regarding the generation of eicosanoids and their impact in inflammation. Lipoxins (LX) are trihydroxytetraene-containing eicosanoids that can be generated within the vascular lumen during platelet-leukocyte interactions and at mucosal surfaces via leukocyte-epithelial cell interactions. During these cell-cell interactions, transcellular biosynthetic pathways are used as major LX biosynthetic routes, and thus, in humans, LX are formed in vivo during multi-cellular responses such as inflammation, atherosclerosis, and in asthma. This branch of the eicosanoid cascade generates specific tetraene-containing products that serve as stop signals, in that they regulate key steps in leukocyte trafficking and prevent leukocyte-mediated acute tissue injury. Of interest here are recent results indicating that aspirin’s mechanism of action also involves the triggering of novel carbon 15 epimers of LX or 15-epi-LX that mimic the bioactions of native LX. Here, an overview of these recent developments is presented, with a focus on the cellular and molecular interactions of these novel antiinflammatory lipid mediators.

Keywords: antiinflammation; lipids; leukocytes; signal transduction; resolution mediators

49z300177
Evasion of Host Immune Surveillance by Hepatitis C Virus: Potential Roles in Viral Persistence
JONATHAN P. MOORMAN (1)(2), MYUNGSOO JOO (2) and YOUNG S. HAHN (2) – (1) Divisions of Geographic Medicine and Infectious Diseases, University of Virginia School of Medicine, Box 485, Charlottesville, Virginia 22908, USA, (2) Beirne B. Carter Center for Immunology Research, University of Virginia Health Sciences Center, Box MR4-4021, Charlottesville, Virginia 22908, USA

Abstract. Hepatitis C virus (HCV) is a major human pathogen that causes mild to severe liver disease worldwide. This positive strand RNA virus is remarkably efficient at establishing chronic infections. In order for a noncytopathic virus such as HCV to persist, the virus must escape immune recognition or evade host immune surveillance. Immune escape via the hypervariable region of the E2 envelope protein has been postulated as one mechanism for HCV persistent infection. Such hypervariability within the E2 protein may be under selective pressure from protective B cell or T cell responses and be able to escape immune recognition by rapid mutation of antigenic site. In addition to antigenic variation, HCV may also suppress immune response, leading to dampening of cellular immunity. This is supported by recent studies in our laboratory demonstrating that the HCV core protein can suppress host immune responses to vaccinia virus by downregulating viral specific cytotoxic T lymphocyte (CTL) responses and cytokine production. An understanding of the mechanisms behind HCV persistence will provide a basis for the rational design of vaccines and novel therapeutic agents targeting human HCV infection.

Keywords: hepatitis C virus; persistence; viral quasispecies; immune evasion

49z301
Peptide Loading of Nascent MHC Class I Molecules
STANISLAV VUKMANOVIĆ, MIRJANA LILIĆ, FABIO R. SANTORI, SANDRA DEMARIA and KIMARY KULIG (Michael Heidelberger Division of Immunology, Department of Pathology, and Kaplan Coprehensive Cancer Center, NYU School of Medicine, 550 First Avenue, New York, NY 10016, USA)

Abstract. A critical molecular interaction during assembly of the major histocompatibility complex (MHC) class I molecules takes place between the heavy chain and the transporter-associated with antigen-processing (TAP) complex. The recent mapping of regions of the heavy chain involved in the binding to TAP suggests a complex molecular interaction essential for the cell surface expression of the MHC class I. The advances made in understanding the TAP-MHC class I interaction are reviewed and discussed here.

Keywords: MHC class I; antigen processing; TAP; CD8+ cells; cytotoxic T lymphocytes.

49z302
Human Antibody Expression in Transgenic Mice
MARIANNE BRÜGGEMANN (Laboratory of Developmental Immunology, The Babraham Institute, Babraham, Cambridge CB2 4AT, UK)

Abstract. Human antibody repertoires can be created in transgenic mice following the introduction of human immunoglobulin heavy and light chain genes in their germline configuration. Transgene constructs or transloci have been obtained by plasmid assembly, cloning in yeast artificial chromosomes, and the use of chromosome fragments. Translocus integration and maintenance in transgenic mouse strains has been achieved by pronuclear DNA injection into oocytes and various transfection methods using embryonic stem cells. The human DNA segments rearrange faithfully in the mouse and produce extensive V(D)J combinations. Specific human monoclonal antibodies of high affinity for use in therapeutic applications have been produced from these translocus mice.

Keywords: human antibody repertoires; transgenesis; human monoclonal antibodies; embryonic stem cell manipulation

49z303
The Search for Novel Adjuvants for Early Life Vaccinations: Can “Danger” Motifs Show Us the Way?
JIRI KOVARIK and CLAIRE-ANNE SIEGRIST (World Health Organization Collaborating Centre for Neonatal Vaccinology, Departments of Pathology and Pediatrics, University of Geneva Medical School, Geneva, Switzerland)

Abstract. Potent but safe adjuvants are required to circumvent the many limitations of the newborn immune system to induce rapidly effective and long lasting immunity to subunit vaccines. By the use of pattern recognition receptors, antigen-presenting cells (APC) can very efficiently be activated by “danger” motifs expressed by various pathogens. APC activated by “danger” motifs, such as immunostimulatory sequences of bacterial DNA, can not only transmit the activation signal from the innate immunity to the adaptive compartment, but also shape the antigen-specific immune responses. Molecules or compounds expressing “danger” motifs could, therefore, be considered for use as adjuvants for subunit vaccines. In this review, the authors discuss the promises and potential drawbacks that such novel adjuvants could hold for their use in experimental and clinical early life vaccinations.

Keywords: vaccines; adjuvants; newborns; infants

49z304
CD89: the Human Myeloid IgA Fc Receptor
H. CRAIG MORTON and PER BRANDTZAEG (Laboratory for Immunohistochemistry and Immunopathology (LIIPAT), Institute of Pathology, University of Oslo, The National Hospital, Rikshospitalet, 0027 Oslo, Norway)

Abstract. CD89 (FcalphaRI) is the human myeloid IgA Fc receptor expressed on cells, such as neutrophils, eosinophils and monocytes/macrophages. Cross-linking of CD89 on these cells, by IgA-opsonised particles (e.g. bacteria, viruses) or anti-CD89 monoclonal antibodies, can trigger various immunological effector functions which are generally protective but may also cause harm to the body. CD89 is a transmembrane glycoprotein that binds both subclasses of IgA in all its molecular forms (i.e. monomeric, dimeric and secretory IgA) via a region of its membrane-distal EC1 domain. DNA studies have shown that the CD89 gene is located within the newly described leukocyte receptor cluster (LRC) on chromosome 19. CD89 is more closely related to the KIR and MIR proteins, whose genes are also found in the LRC, than to other human Fc receptors (FcRs). On myeloid cells, CD89 is able to associate with the immunoreceptor tyrosine-based activation motif (ITAM)-containing the FcR gamma chain, which is responsible for intracellular signaling via CD89. Recently, it has been suggested that some cells express CD89 in a form that does not associate with the FcR gamma chain. Although the biological relevance of this observation is not yet clear, it may explain certain anti-inflammatory/inhibitory effects attributed to IgA. Here we review current knowledge concerning the genetics, structure and biological function of CD89.

Keywords: CD89; FcalphaR; FcalphaRI; IgA; Fc receptor; myeloid

49z305
Immune-Endocrine Interactions of the Hypothalamus-Pituitary-Thyroid Axis: Integration, Communication and Homeostasis
MATTHEW D. ARMSTRONG and JOHN R. KLEIN (Department of Biological Science and the Mervin Bovaird Center for Studies in Molecular Biology and Biotechnology, University of Tulsa, Tulsa, OK, USA)

Abstract. The immune and neuroendocrine systems are two essential physiological components of mammalian organisms. Although each is primarily committed to a set of tasks involved, on the one hand, in the protection from infection and disease, and on the other hand, in the regulation of metabolism and other physiological activities, there is also evidence indicating that active and dynamic collaborations exist between those systems in the execution of their designated functions. These interactions occur at many stages of embryonic and neonatal development, and they are a continual part of the normal homeostatic balance needed to maintain health. The present review discusses various historical and contemporary perspectives of immune-endocrine interactions involving the hypothalamus-pituitary-thyroid axis, and offers a hypothesis of how this aspect of the neuroendocrine system participates directly in the immune response to antigenic challenge, infection and disease.

Keywords: immune-endocrine; hormone; antigen-presenting cells; pituitary-thyroid; lymphocytes; immunity

49z306

Cellular Immunology

Involvement of Beta2-Microglobulin in CD69 Expression on T Cells
LESZEK PĄCZEK, BOŻENA CZARKOWSKA-PĄCZEK, GRAŻYNA KORCZAK-KOWALSKA, PIOTR WIERZBICKI, IRENA BARTŁOMIEJCZYK and ANDRZEJ GÓRSKI (The Transplantation Institute, Medical University of Warsaw, Nowogrodzka 59, 02-006 Warsaw, Poland)

Abstract. Beta2-Microglobulin (beta2M) is the light chain of the class I HLA molecule. The serum level of beta2M is elevated in various diseases including lymphoma, inflammation, viral infections and chronic renal dysfunction. The present study addressed the possible influence of beta2M on T lymphocyte activation in vitro. Peripheral blood mononuclear cells from a group of 17 healthy subjects were examined. Stimulation with OKT3 and fibronectin in combination with 30 mg beta2M/dl resulted in a two-fold increase of cell proliferation. A similar effect was observed when OKT3 and collagen I were applied as well as when OKT3 and collagen IV were used as costimulation to T cells. The CD69 expression, measured by flow cytometry was significantly enhanced above the control level (1.52 ± 1.03% vs 33.21 ± 20.26%, p<<0.01, control group and 30 mg beta2M/dl, respectively). Together, these observations suggest that beta2M may play a role in modulating lymphocyte proliferation, possibly through modification of the CD69 molecule.

Keywords: beta2-microglobulin; mononuclear cells; proliferation; CD69

49z307
In Vitro and in Vivo Study of the Expression Vector Encoding Vascular Endothelial Growth Factor
MACIEJ MAŁECKI, MAŁGORZATA PRZYBYSZEWSKA and PRZEMYSŁAW JANIK (Department of Cell Biology, Cancer Center, Roentgena 5, 02-781 Warsaw, Poland)

Abstract. Vascular endothelial growth factor (VEGF) is an angiogenic cytokine with potential therapeutic applications in human diseases. It is a mitogen primarily for endothelial cells. The transfer of the cDNA encoding VEGF to ischemic tissues, which cannot be revascularized otherwise, represents a novel and promising approach to the treatment of vascular disorders. In this work the VEGF165 cDNA was cloned into the expression vector pSecTag2B. The activity of the construct was studied in cell culture as well as in vivo. Western blotting study showed that the cells transfected with the vector secreted significantly higher amounts of VEGF to the culture medium than the non-transfected cells. In vivo study revealed an increased number of new vessels in animals injected with vector encoding VEGF as compared with empty plasmid. Also, tumor cells transfected with the VEGF plasmid exhibited extensive vascularization.

Keywords: angiogenesis; VEGF; gene therapy

49z308

Various

Modification of Collagen Film by Certain Chemical Agents
EWA LESIAK-CYGANOWSKA, DARIUSZ ŚLADOWSKI and JANUSZ KOMENDER (Department of Transplantology and Central Tissue Bank, Center of Biostructure Research, Warsaw Medical University, Chałubińskiego 5, 02-004 Warsaw, Poland)

Abstract. Morphological changes and the content of free carboxyl groups in bovine collagen (type I) film under the influence of trypsin, hydrochloric acid (HCl) and ethylenediaminetetraacetic acid (EDTA) were studied. Incubation with trypsin and HCl was found to cause some delamination of the film and the appearance of some low-density spots. Incubation with EDTA did not cause any morphological changes. A high concentration of free carboxyl groups (10-fold higher than in control) was seen after incubation with trypsin.

Keywords: collagen modification

49z309
In Vitro Culture of Human Epithelial Cells on a Modified Xenogenic Collagen Support
EWA LESIAK-CYGANOWSKA (1), DARIUSZ ŚLADOWSKI (1), EWA JANKOWSKA (2) and JANUSZ KOMENDER (1) – (1) Department of Transplantology and Central Tissue Bank, Centre of Biostructure Research, Warsaw Medical University, Chałubińskiego 5, 02-004 Warsaw, Poland, (2) Department of Electron Microscopy, Centre of Biostructure Research, Warsaw Medical University, Chałubińskiego 5, 02-004 Warsaw, Poland

Abstract. Human epithelial cells (HeLa, HaCaT, NHK) were cultured in vitro on chemically modified collagen membranes. Adhesion to the support was measured by estimation of the percentage of adhering 51Cr-labeled cells. Proliferation was estimated with the XTT test. Morphological observations of cells growing on HCl-treated collagen were performed using histological and electron microscopic techniques. HCl and trypsin-modified xenogenic collagen was found to be a good support for human cells in vitro. EDTA-incubated collagen enhanced neither adhesion nor proliferation. The best adhesion and proliferation were found on HCl-treated collagen, depending, however, on the kind of cells.

Keywords: collagen modification; keratinocytes growth

49z310

Vol. 49, No. 4, 2001

Review

Antigen Receptor Signaling Is Subverted by an Immunomodulatory Product Secreted by a Filarial Nematode
MARGARET M. HARNETT (Departments of Immunology, University of Glasgow, Glasgow G11 6NT, UK) and WILLIAM HARNETT (University of Strathclyde, Glasgow G4 ONR, UK)

Abstract. ES-62 is a phosphorylcholine (PC)-containing glycoprotein secreted by the rodent filarial nematode Acanthocheilonema viteae which is able to inhibit antigen receptor-stimulated proliferation of B and T lymphocytes in vitro and in vivo. The active component of ES-62 appears to be PC as the results obtained with ES-62 are broadly mimicked by PC conjugated to bovine serum albumin or PC alone. Such desensitization of lymphocyte responsiveness appears to reflect an uncoupling of the antigen receptors from key intracellular proliferative signaling events, such as the phosphoinositide-3-kinase (PI-3K), protein kinase C (PKC) and Ras mitogen-activating protein kinase (RasMAPK) pathways. ES-62 mediates such immunomodulatory effects at concentrations equivalent to those found for PC-containing molecules in the bloodstream of parasitized humans and, thus, ES-62 provides a model system for dissecting the mechanisms of immune evasion induced by related PC-containing glycoproteins expressed by human filarial nematodes.

Keywords: filarial nematode; immune evasion; lymphocyte; phosphorylcholine; signal transduction

49z401
Enhancing Cytotoxic T Cell Responses with Altered-Peptide Ligands
RUI ZHAO and EDWARD J. COLLINS (Department of Microbiology and Immunology, and Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599)

Abstract. Interest in class I MHC-mediated immunotherapy is growing rapidly. In order to fight a virus or cancer effectively, a successful immunotherapeutic must activate a large number of specific CD8+ T cells and also generate immunological memory. Attempts to generate immune responses towards tumor- or virus-derived peptides have frequently been frustrated by the nature of the peptide antigen itself. Either the peptide does not bind well to its cognate MHC, or the T cells directed towards it have been functionally inactivated in vivo. Altered-peptide ligands are an effective way to circumvent these problems. However, generating enhanced binding of altered peptides to class I MHC while still maintaining recognition of the wild-type peptide is not straightforward. Many groups design enhanced binding peptides by substituting the observed anchor residues with those that are most preferred by the class I MHC molecule. For many antigenic peptides, this approach does not work. Furthermore, if a higher affinity peptide is designed, the substitutions may result in reduced recognition by CD8+ T cells. Therefore, the design of an altered-peptide ligand requires careful testing of each candidate therapeutic in terms of affinity for class I MHC and immunological reactivity. Lastly, immunotherapy using class I MHC must also take into account the large genetic heterogeneity in the population. A therapeutic that is only effective for 5-10 percent of the population is not as attractive as one that works for over 90% of the population. The use of MHC supertypes (groups of class I MHC allotypes that share similar peptide-binding characteristics) shows great promise in overcoming this problem.

Keywords: class I MHC, T cell receptor, ligand design, immunotherapy, x-ray crystallography

49z402
Multi-Functional Roles of Stat3 Revealed by Conditional Gene Targeting
KIYOSHI TAKEDA and SHIZUO AKIRA (Department of Host Defense, Research Institute for Microbial Diseases, and CRSET of Japan Science and Technology Corporation, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan)

Abstract. Signal transducer and activator of transcription (STAT) is a family of transcription factors composed of seven members. Gene-targeted mice of each STAT family protein displayed defective responses to cytokines, demonstrating an important role in cytokine-mediated biological responses. However, unlike the mice lacking other STAT proteins, Stat3-deficient mice died during their early embryogenesis. Therefore, in an attempt to avoid the lethality and assess the role of Stat3 in cytokine-mediated functions in mouse adult tissues, conditional gene targeting utilizing a Cre-loxP system was achieved. By this method, Stat3 was disrupted in several types of tissue, including T cells, macrophages, skin, and mammary gland. Analyses of these Stat3-mutant mice revealed important roles of Stat3 in biological functions in each tissue.

Keywords: Stat3, conditional gene targeting, cytokine, signal transduction

49z403
Redox Control of Cellular Function by Thioredoxin; a New Therapeutic Direction in Host Defence
YUMIKO NISHINAKA, HAJIME NAKAMURA, HIROSHI MASUTANI and JUNJI YODOI (Department of Biological Responses, Institute for Virus Research, Kyoto University, 53 Shogoin-Kawaharacho, Sakyo, Kyoto 606-8507, Japan)

Abstract. Compelling evidence has suggested that oxidative stress mediates various cellular reponses, and control of reduction/oxidation (redox) is importan in maintaining the homeostatsis of an organism. The thioredoxin (TRX) system, along with as well as the glutathione system, is one of the key system in controling cellular redox statuts. TRX is a small ubiquitous protein with the redox-active site sequence -Cys-Gly-Pro-Cys-. It has been demonstrated to be a multifunctional protein, which has regulatory roles in cellular signaling and gene transcription in addition to cytoprotective activities through the quenching of reactive oxygen species. Various oxidative stimuli, such as as UV irradiation, cytokines and some chemicals, promptly induce the xpression of TRX. Overexpression of TRX correlates with a wide variety of oxidative stress conditions and, in some cases, TRX has shown promising effects for clinical use, for instance in the attenuation of tissue injury in ischemia reperfusion models. The modulation of TRX functions in association with other redox-regulatory should give us a new therapeutic strategy in the treatment of oxidative stress-mediated disorders and diseases.

Keywords: redox regulation, thoredoxin, reactive oxygen species, therapy

49z404
Cytokine-Based Immunotherapy of Allergic Disease
IAN P. LEWKOWICH and KENT T HAYGLASS (Department of Immunology, University of Manitoba, Winnipeg, Canada)

Abstract. Human immediate hypersensitivity diseases are strongly associated with an excessive type 2 response to normally innocuous environmental antigens, and are a growing health care concern in developed nations. Commonly prescribed treatments provide effective symptomatic relief, but are unable to consistently ameliorate the underlying cause of allergic disease: the excessive generation of allergen specific Th2 cells. IL-12 and IL-18 are potent inducers of type 1 immunity, and, as such, have been proposed as candidates for treatment of allergic diseases. This review critically assesses the potential of recombinant IL-12 and IL-18 immunotherapy to redirect both de novo and established allergic responses in animal models of human allergic disease to clinically protective immune responses.

Keywords: IL-12, IL-18, immunotherapy, immediate hypersensitivity

49z405
Evidence for an Immunoregulatory Role of OX2 with Its Counter Ligand (OX2L) in the Regulation of Transplant Rejection, Fetal Loss, Autoimmunity and Tumor Growth
REG M. GORCZYNSKI (CCRW 2-855, The Toronto Hospital, University Health Network, Toronto, Canada, and Departments of Surgery and Immunology, University of Toronto, Toronto, Canada)

Abstract. Transplantation has emerged as an effective treatment for patients with end-stage organ failure. Current regimens of non-specific immunosuppressive drug treatment, which are needed life-long to prevent graft rejection, have numerous adverse side effects and increase the risk of opportunistic infections and malignancy. A major goal is to develop immunotherapeutic protocols that achieve specific tolerance. Such protocols would decrease and eventually eliminate the reliance on non-specific drug therapy. We showed that portal vein (pv) delivery of donor antigen prolongs the survival of vascularized and non-vascularized allo- and xeno-grafts, and that increased graft survival is associated with altered cytokine production and augmented expression of the molecule OX2. This review documents further evidence for a more general immunoregulatory role for the interactions of OX2 and its ligand, OX2L.

Keywords: OX2; immunoregulation; tolerance; transplantation; autoimmunity; fetal loss syndrome

49z406

Clinical Immunology

The Levels of IL-1beta, IL-4 and IL-6 in the Serum and the Liver Tissue of Chronic HCV-Infected Patients
TADEUSZ WOJCIECH ŁAPIŃSKI (Department of Infectious Diseases, Medical Academy of Białystok, Żurawia 14, 15-540 Białystok, Poland)

Abstract. The pro-inflammatory interleukines play a major role in the progress of chronic hepatitis C. Among the patients with chronic HCV infection, the morphology of the liver was assessed and the levels of serum and liver-tissue IL-1beta, IL-4 and IL-6 were determined. The levels of the cytokines were related to the liver tissue changes. RNA-HCV was measured by the RT-PCR method. Cytokine levels of the serum and liver tissue were measured by the Quantikine High Sensitivity test. The levels of serum IL-1beta, IL-4 and IL-6 (0.221, 0.104 and 1.393 pg/ml) in all HCV patients were higher in comparison with healthy adults (0.188, 0.025 and 0.600 pg/ml). The levels of liver tissue IL-1beta, IL-4 and IL-6 (4291.3, p<0.05; 1624.6, p<0.05; 1158.7 pg/g protein) in all HCV patients were higher compared to the patients with liver cirrhosis without HBV or HCV infection (2319.9, 553.6 and 756.2 pg/g protein). Patients with HCV infection demonstrated significant correlation between serum and liver-tissue levels of IL-1beta (Pearson: 0.61, p<0.05) and IL-4 (Pearson: 0.51). The level of serum IL-6 in patients with moderate chronic active hepatitis was higher when compared to the patients with mild chronic persistent hepatitis. Among the patients with mild chronic persistent hepatitis, the levels of liver tissue IL-6 were higher compared with those with moderate chronic active hepatitis. There was no correlation between histology changes and the levels of serum and liver-tissue IL-1beta and IL-4.

Keywords: HCV infection, cytokines, histology changes

49z407
Soluble Selectin Profiles Associated with Severe Trauma
ANDRZEJ SIEMIĄTKOWSKI (Department of Anesthesiology and Intensive Therapy), FRANCISZEK ROGOWSKI (Department of Nuclear Medicine), URSZULA WERESZCZYŃSKA-SIEMIĄTKOWSKA (Department of Gastroenterology), LIDIA MALINOWSKA (Department of Anesthesiology and Intensive Therapy) and JACEK BORKOWSKI (Department of Anesthesiology and Intensive Therapy, Medical Academy of Białystok, M. Skłodowskiej-Curie 24a, 15-276 Białystok, Poland)

Abstract. Severe trauma acts as a trigger for the complex cascade of postinjury events leading to the release of different mediators and the development of generalized inflammation. Selectins are a family of adhesion proteins that are responsible for the adherence of polymorphonuclear neutrophils to the endothelium. This interaction plays an important role in the development of severe complications after multiple trauma. The aim of the present study is to follow the sequential alterations in circulating selectins level after severe injury and to evaluate the clinical significance of these mediators in monitoring prognosis and outcome. Thirty four severely traumatized patients were entered into the study. Serum sE-selectin, plasma sP-selectin and sL-selectin concentrations were measured and an APACHE II score was calculated on admission to the intensive care unit and during the subsequent 5 days. The patients were divided into survivors and nonsurvivors. Initial soluble P- and E-selectin concentrations were significantly elevated in all trauma patients. The highest values of these adhesion molecules were measured in all the observed days in patients with poor prognosis and outcome. In survivors we found a systematic decrease in the sP-selectin concentrations. On admission, the sL-selectin concentrations in all trauma patients were decreased. There were stable, very low values in nonsurvivors and a slow increase in circulating L-selectin in patients who survived. The pattern of soluble selectins in patients with severe trauma is characterized by increased levels of P- and E-selectin and a decreased concentration of L-selectin. These findings suggest a widespread microvascular endothelial activation on injury in the early posttraumatic period, which may be associated with increased neutrophil – endothelial adhesion, neutrophil extravasation and migration. We suppose that these parameters of endothelial cell activation/injury may be useful as another early prognostic factor in severe trauma.

Keywords: severe trauma, endothelium, selectins, prognosis

49z408
Lactoferrin Regulates the Immune Responses in Post-Surgical Patients
MICHAŁ ZIMECKI (Department of Experimental Therapy, Institute of Immunology and Experimental Therapy, Polish Academy of Science, Weigla 12, 53-114 Wrocław, Poland), ANDRZEJ WŁASZCZYK (Department of Anesthesiology and Intensive Therapy Wrocław Academy of Medicine, Chałubińskiego 1a, 50-368 Wrocław, Poland), ROBERT WOJCIECHOWSKI (Department of Anesthesiology and Intensive Therapy Wrocław Academy of Medicine, Chałubińskiego 1a, 50-368 Wrocław, Poland), JANUSZ DAWISKIBA (First Clinic of General and Endocrine Surgery, Wrocław Academy of Medicine, Poniatowskiego 2, 50-326 Wrocław, Poland) and MARIAN KRUZEL (Department of Integrative Biology, Pharmacology and Physiology, University of Texas, Medical School at Houston Texas, USA)

Abstract. The effect of oral administration of lactoferrin (LF) was studied to determine if it could modify post-surgical immune response. The action of lactoferrin was evaluated in 18 LF-treated patients versus 28 placebo counterparts. Patients (women and men, mean age 50 years) were given daily oral doses (20 mg each) of LF for 5 consecutive days prior to thyroid surgery. The following immune response parameters were determined in blood samples taken from the patients one day before, one day after, and 5-7 days following surgery: cell morphology, the proliferative response of peripheral blood mononuclear cells (PBMC) to phytohemagglutinin (PHA), and the spontaneous and lipopolysaccharide (LPS)-induced production of tumor necrosis factor alpha (TNF-a) and interleukin 6 (IL-6). As a consequence of the thyroid surgery, the total leukocyte count increased on the postoperative day by about 50% in all patients and the percentage of lymphocytes fell by 26 and 35% in the control vs LF-treated group. The content of neutrophils, on the other hand, elevated on day 1 post-operation by 51 and 68%, respectively. The percent of neutrophil precursors was markedly higher in LF-treated patients, particularly on the day before and the day after surgery (4.1 and 4.8 vs 2.5 and 3.7%, respectively). The post-surgical values were, however, comparable in both groups for neutrophils. The proliferative response of lymphocytes showed a slight decrease in the control group and an increase in the LF-treated patients on day 5 post-operation (20% over control group). LPS-induced TNF-a production was higher in LF-treated patients in both one day before and one day following surgery (28 and 24% respectively). LPS-induced IL-6 production was comparable in both placebo and LF-treated patients before surgery, however, on day 1 and 5 following surgery, the production of IL-6 was higher in LF-treated patients by 65 and 27%, respectively. Taken together, the data presented in this study revealed increased immune responsiveness in all patients treated with lactoferrin subjected to the thyroid surgery. This suggests that treatment with lactoferrin could constitute an effective protective measure against post-surgical complications.

Keywords: lactoferrin, clinical insult, immunoregulation, prevention

49z409

Vol. 49, No. 5, 2001

Review

Cancer Immunogene Therapy
Hirohisa Yoshizawa, Hiroshi Kagamu and Fumitake Gejyo (Department of Medicine (II), Niigata University Medical School, Niigata, Japan)

Abstract. The establishment of cancer in a host involves at least two major events: the escape of tumor cells from normal growth control and their escape from immunological recognition. Because of this nature of their development, cancer cells seem to be predominatly poorly immunogenic. In contrast to the previous idea that cancer cells express no recognizable antigens, recent progress in the identification and characterization of tumor antigens, as well as the expansion of knowledge on the cellular and molecular mechanisms of antigen recognition by the immune system, have raised the possibility of using immunotherapy to treat certain tumors. Information on these mechanisms has been obtained in three crucial areas: 1) the role of cytokines in the regulation of the immune response, 2) the molecular characterization of tumor antigens in both mouse and human tumors, and 3) the molecular mechanisms of T cell activation and antigen presentation. Such information has provided new insight into tumor immunology and immunotherapy. Furthermore, recombinant DNA technology allows for modification of the genome of mammalian cells for therapeutic purposes in several diseases. Several novel strategies have been developed to derive genetically modified tumor cells and use them as cellular vaccines to induce antitumor immunity in animal tumor models. This combined modality of genetically modified tumor cells and immunotherapy has been termed immunogene therapy of tumors. Crucial to this approach has been the ability to transfer into normal or neoplastic cells genes known to increase the immunogenicity of cells, which subsequently can be used to augment immune reactions in tumor-bearing mice or cancer patients. While there has been success in inducing antitumor immunity in some tumor models, there are difficulties and limitations in the application of these gene-modified tumor cells for the treatment of preexisting tumors. In this review, recent progress in cancer immunogene therapy is discussed.

Keywords: immunotherapy; transfection; cytokine; costimulatory molecule.

49z501
Regulation of IL-5 expression
Viatcheslav A. Mordvinov and Colin J. Sanderson (Curtin University of Technology, Perth, Western Australia)

Abstract. Interleukin-5 (IL-5) is a cytokine primarily involved in the pathogenesis of atopic diseases. It specifically controls the production, activation and localization of eosinophils, the major cause of tissue damage in atopic diseases. IL-5 belongs to a gene family shared by IL-3, IL-4 and GM-CSF and is predominantly regulated at the transcriptional level. A variety of stimuli and modulators have been identified as regulating production of IL-5 both in vivo and in vitro, indicating a highly complex series of control mechanisms. However, a better understanding of the biology of IL-5 and the regulation of its expression is crucial for the development of new therapeutic agents for allergic disease. This review covers the major molecular aspects of IL-5 research.

Keywords: interleukin 5; cytokines; allergic diseases.

49z502
Sjogrens’s Syndrome: Imunological Response Underlying the Disease
Jason B. Brayer, Michael G. Humphreys-Beyer and Ammon B. Peck (Departments of Oral Biology and Pathology, Laboratory Science and Center for Orphaned Autoimmune Diseases, University of Florida, Gainesville, FL 32610, USA)

Abstract. Sjögren’s syndrome is a chronic autoimmnune disorder characterized primarily by the discomforts od dry eyes and dry mouth due to the progressive loss of exocrine gland function. Development of a number of animal models to study Sjögren’s syndrome, especially the NOD mouse and its congenic partner strains, has permitted a systematic analysis of immunological and non-immunological factors that influence predisposition for development of the autoimmune response. These data are reviewed here.

Keywords: NOD mouse; autoimmune exocrinopathy; cytokine knock-out mice; autoantibody.

49z503
B Cell Tolerance to Self in Systemic Autoimmunity
Moncef Zouali (Hôpital Broussais, INSERM U 430, 96 rue Didot, 75674 Paris Cedex 14, France)

Abstract. After a century of research and despite intensive scrutiny, the origin of autoantibody production remains an enigma. Recently, the essential role of B cells in promoting systemic autoimmunity in mice seems more important than previously thought: self-reactive B cells can be subject to positive selection and a deficiency in serum IgM predisposes to the development of IgG antibodies to autoantigens. Studies of the B cell repertoire expressed in systemic autoimmune diseases have provided important clues. In human lupus, quantitation of this repertoire reveals the presence of an expansion of IgG clonotypes that impart reactivity with disease-related autoantigens. The nucleotide sequences of autoantibodies derived from these patients and expressing nephritogenic idiotopes (present in immune complexes and renal eluates of subjects with active disease) show features of diversification with a high rate of replacement/silent mutations and clustering of the mutations in the hypervariable regions, suggesting than an antigen-driven process plays a role in the generation of pathogenic autoantibodies. Currently, the contributions of apoptosis and of cell receptor signaling to this triggering are being appreciated. Pursuing these and related issues will have an important impact on autoimmune research.

Keywords: autoimmunity; B cells; systemic lupus erythematosus.

49z504
HIV-Macrophage Interactions at the Cellular and Molecular Level
Makobetsa Khati, William James and Siamon Gordon (Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom)

Abstract. Macrophages, centrally involved in both the innate and adaptive arms of the immune system, are not only the chief target of the human immunodeficiency virus (HIV), but also its main reservoir and vehicle of transmission. Macrophage-tropic viruses are responsible for the initial infection, predominate in the asymptomatic phase, and persist throughout infection, even after the emergence of preferential T cell- and/or dual-tropic HIV-1 variants. Functional impairment of HIV-infected macrophages plays a role in the immune dysregulation characteristic of AIDS (acquired immunodeficiency syndrome). Efforts directed ant understanding the cellular and molecular mechanisms underlying HIV-macrophage interactions remain the basis for devising novel and efficacious therapeutic strategies against HIV and the AIDS epidemic.

Keywords: macrophages; HIV reservoir; immune dysregulation; AIDS.

49z505
Mycobacterium-Specific Human CD8 T Cell Responses
Michel R. Klein and Annette Fox (Tuberculosis Research Programme, Medical Research Council (MRC) Laboratories, Atlantic Road, Fajara, PO Box 273, Banjul, The Gambia, West Africa)

Abstract. Tuberculosis (TB) remains a global health problem. There is an intense effort to identify correlates of protective immunity and to design new TB vaccines. CD8 T cells are thought to play a significant role in controlling Mycobacterium tuberculosis infection. Relatively little has been published about the antigens and epitopes targeted by mycobacteria-specific CD8 T cells. Here we present an update of our 1999 overview of human CD8 T cell epitopes in mycobacterial antigens and discuss related issues relevant to TB diagnosis and vaccine development.

Keywords: mycobacterium; tuberculosis; CD8; epitope; vaccine; diagnosis.

49z506

Clinical Immunology

The Effects of Endotoxin Administration on Cytokine Production in Obstructive Jaundiced Rats
Janusz Dawiskiba¹, Michał Zimecki², Danuta Kwiatkowska³ and Adam Kuźmiński¹

Abstract.
¹First Clinic of General and Endocrine Surgery, Wrocław Academy of Medicine, Poniatowskiego 2, 50-326 Wrocław, Poland,
²Department of Experimental Therapy, Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Weigla 12, 53-114 Wrocław, Poland,
³Department of Clinical Biochemistry, Wrocław Academy of Medicine, Chałubińskiego 10, 50-368 Wrocław, Poland

The aim of this study was to evaluate the effect of lipopolysaccharide (LPS) administration, which mimics a surgical intervention, on the immune status of obstructive jaundiced (OJ) and sham-operated control rats. Rats were given 20 mg LPS intraperitoneally on day 13 following bile duct ligation or sham surgery. We determined serum levels of tumor necrosis factor a (TNF-a) and interleukin 6 (IL-6) on day 14 after surgery, and spontaneous as well as LPS-induced production of these cytokines in splenocyte and peritoneal exudate cell cultures (PEC). We found that IL-6, but not TNF-a, serum concentrations were significantly elevated (4-fold) in OJ rats treated with LPS compared with LPS-untreated OJ rats. In sham-operated rats the differences between the respective groups were not significant. The production of TNF-a by splenocyte and PEC cultures was depressed in OJ rats treated with LPS; in particular, a very deep decline was observed in the case of spontaneous TNF-a production in PEC cultures. In contrast, TNF-a production in LPS-untreated and LPS-treated sham-operated rats did not differ. In the case of IL-6 production by splenocytes and PEC cultures, we observed a significant suppression of this cellular function in both OJ and sham-operated rats treated with LPS when compared with the respective controls. In conclusion, the results indicate that the already depressed cytokine production in OJ rats leads to even deeper hyporeactivity following LPS challenge. Lack of TNF-a suppression upon LPS treatment in sham-operated rats suggests that surgery-elicited hyporeactivity is mediated by a different mechanism than that leading to immune hyporesponsiveness in OJ. Our findings may explain the relatively high mortality rates of OJ patients subjected to surgery.

Keywords: obstructive jaundice; LPS; splenocytes; peritoneal cells; TNF-a; IL-6.

49z507
Proinflammatory Cytokines (IL-6, IL-8), Cytokine Ihibitors (IL-6 sR), sTNFR II) and Anti-Inflammatory Cytokines (IL-10, IL13) in the Pathogenesis of Sepsis in Newborns and Infants
Janusz Piotr Sikora, Danuta Chlebna-Sokół (Department of Pediatric Propedeutics, Institute of Pediatrics, Medical University of Łódź, Poland) and Alicja Krzyżańska-Oberbek (Critical Care Unit, District Hospital No. 4, Medical University of Łódź, Poland)

Abstract. The levels of the proinflammatory cytokines interleukin 6 (IL-6) and IL-8, and the anti-inflammatory cytokines IL-10 and IL-13 were studied in child patients with sepsis. The changes of the cytokines inhibitors soluble IL-6 receptor and soluble p75 TNF alpha receptor were also investigated in the patients’ sera. An increase of pro- and anti-inflammatory cytokine levels was demonstrated at the time of diagnosis. Pharmacotherpy was accompanied by a decrease of the elevated concentrations of both cytokines and their inhibitors. The time pattern of changes in cytokine and cytokine inhibitor serum concentrations along with the time course of acute phase indices, including procalcitonin and C-reactive protein, allows for an evaluation of system inflammatory response and may support diagnostic and prognosis methods.

Keywords: sepsis; cytokines; cytokine inhibitors; children.

49z508

Vol. 49, No. 6, 2001

Review

Mechanisms of Mouse T Lymphocyte-Induced Suppression of the IgG_2a^b Allotype and T Lymphocyte Tolerance to IgG_2a^b.
Lalech Majlessi and Guy Bordenave

Abstract. In mice of the Igh^aimmunoglobulin allotypic haplotype we found, the presence of T lymphocytes with an inherent inhibitory activity against the expression of the IgG_2a^b allotype (IgG_2a of the Igh^b immunoglobulin allotypic haplotype). This constitutive anti-IgG_2a^b T lymphocyte activity can be enhanced in vivo by what we called „sensitization”, which usually consists of one or two intravenous injections of B splenocytes from Igh^b congenic mice. When injected at birth, the resulting anti-IgG_2a^b T splenocytes induce, with 100% success, total, specific and chronic (but experimentally reversible) suppression of IgG_2a^b in Igh^a/b F₁ hybrid mice prepared by mating Igh congenic mice. Even if restricted to IgG_2a^b expression, this experimental model, which deals with an unambiguous case of T cell-mediated down-regulation of immu oglobulin production, provides a clear and powerful tool to dissect finely the behavior of the partners(T and B lymphocytes) intervening in regulation within the immune system. For example, we observed that CD4 T lymphocytes were necessary to obtain full recruitment of anti-IgG_2a^b CD8 T lymphocytes during the sensitization, that suppression induction in anti-IgG_2a^b T splenocytes of newborn recipients required cooperation between CD4 and CD8 T lymphocytes, and that CD8 T lymphocytes were essential for suppression maintenance. We showed that this suppression was not characterized by an accumulation of B lymphocytes containing the allotype they could not secrete or Cg_2a^b mRNA they could not translate. The recipient’s immune system was not involved in the suppession maintenance; this was done by donor T lymphocytes, which ensured the chronicity of IgG_2a^b suppression throughout the recipient’s life. We demonstrated that the mechanism of this suppression implied an MHC-restricted presentation by target B lymphocytes of Cg_2a^b peptides to the T cell receptor (TCR) of anti-IgG_2a^b T lymphocytes. Notwithstanding the requirement of a CD4-CD8 T lymphocyte cooperation during the induction phase, we functionally determined that the suppression induction implicated an MHC class I-, but not class II-restricted interaction. We also demonstrated the existence in vivo of alternative or concomitant use of perforine- and Fas-mediated cytotoxicity pathways in this T cell-induced IgG_2a^b suppression. Thus this suppression did not imply silencing IgG_2a^b production, but B lymphocyte destruction by CD8 T lymphocytes. Always using our suppression model, we demonstrated that an agonistic anti-CD40 treatment helps in recruiting CD8 cytotoxic T lymphocytes, involved in immune regulatory functions and that CD40 expression on Igh^b B lymphocytes confronted with CD8 T lymphocyte effectors only operating via the Fas pathway was involved in the total suppression of IgG_2a^b expression. The selection and maintenance of such normal T cell activity against the IgG_2a^b allotype in mice of different genetic backgrounds remain somewhat enigmatic. Indeed, we did not observe any similar activity against other immunoglobulin allotypes or isotypes. The intestinal flora had no influence on the emergence of this anti-IgG_2a^b T lymphocyte activity, as it was untouched in germ-free Igh^a mice when compared with normal Igh^a mice. More recently, this model offered an opportunity to study problems pertaining to immune tolerance. For instance, we showed that the genetic elements involved in the building of anti-IgG_2a^b TCR were available in Igh^a and Igh^b mice of different genetic backgrounds, but that somatic constraints, namely the perinatal presence of IgG_2a^b, effectively prevented their acquisition, while its absence led to their spontaneous emergence. Consequently, we were able to induce anti-IgG_2a^b T lymphocytes into a tolerance state by injecting Igh^a mice with soluble IgG_2a^b during the perinatal period. However, the full T lymphocyte tolerance obtained in this manner was not definitively acquired, as it had reversed spontaneously when investigated 3 to 6 months after the end of tolerogen treatment, even when this treatment had been prolonged from the perinatal period to 9 months of age. The mechanisms (induction and reversion) of this tolerance involves the physical elimination or the irreversible inactivation of the natural anti-IgG_2a^b T lymphocyte clones and their resurgence, from bone-marrow precursors, as long as the thymus remains operational, but not the establishment of a reversible, functional unresponsiveness (anergy) or an active, cell-mediated inhibition of anti-IgG_2a^b T clones. We attempted to elucidate, in Igh^b mice, whether the natural T lymphocyte unresponsiveness to IgG_2a^b involved a central tolerance mechanism and to identify the type of tolerogen implicated in this tolerogenesis. The experiments principally showed that this natural T lymphocyte tolerance to IgG_2a^b was mediated by a thymic mechanism; that the capacity to induce it was gradually acquired by Igh^b thymuses and was most probably due to potentially IgG_2a^b-producing/presenting cells, progressively colonizing the developing thymus; and that a significantly decreased postnatal Cg_2a^b gene transcription correlated with the emergence of anti-IgG_2a^b T lymphocytes in Igh^a/b F₁ (postnatally deprived of their B lymphocyte compartment), which subjected them to autoimmune IgG_2a^b-allotype suppression.

Keywords: T lymphocytes; IgG_2a^ballotype; suppression; tolerance; cytotoxicity; down-regulation of Ig production.

49z601
Interleukin 10 and Its Role in the Regulation of the Cell-Mediated Immune Response in Syphilis
Marzanna Łusiak and Jadwiga Podwińska

Abstract. Data concerning interleukin 10 (IL-10), a cytokine of Th2 lymphocytes, and its inhibition of Th1 lymphocytes from secreting interleukin 2 (IL-2) and interferon (IFN) are presented. It has been indicated that IL-10 also inhibits other cells from producing interleukin 12 (IL-12) and nitric oxide (NO). It is known that all these factors take part in the cell-mediated immune response and immunity. This inhibition may facilitate the multiplication of Treponema pallidum and the development of disease despite the presence of immunologically competent cells. It has also been demonstrated that in late latent syphilis, when Th1 lymphocytes are not able to produce IL-2 and IFN, the cells are able to produce only IL-12 and NO. This fact seems to suggest that these factors take over the immune function when cells are stimulated again by treponemes which, after many years of latency, begin to multiply. Thus, a high level of IL-12 and NO seems to be an indicator of the development of the third stage of disease.

Keywords: IL-2; IFN; IL-10; IL-12; nitric oxide; cell-mediated immune response; syphilis.

49z602

Clinical Immunology

Serum Autoantibodies Profile and Increased Levels of Circulating Intercellular Adhesion Molecule-1: a Reflection of the Immunologically Mediated Systemic Vasculopathy in Rheumatic Diseases?
Anna Kuryliszyn-Moskal, Piotr Adrian Klimiuk and Stanisław Sierakowski (Department of Rheumatology and Internal Diseases, Medical University of Bialystok, M. Skłodowskiej-Curie 24a, 15-276 Białystok, Poland)

Abstract. Clinical manifestation of systemic vasculitis may be postulated as a consequence of the immune response abnormalities in the course of connective tissue diseases (CTD). The aim of this study was to elucidate the significance of the different autoantibodies and soluble intercellular adhesion molecule 1 (sICAM-1) shedding into the circulation in the diagnosis of vasculitis in rheumatic diseases. Serum of 86 patients with rheumatic diseases (54 with rheumatoid arthritis (RA) and 32 with CTD) were analyzed for the concentrations of sICAM-1 levels by the enzyme linked immunosorbent assay (ELISA). Control sera were obtained from 30 healthy individuals. Anti-nuclear antibodies (ANA), anti-double-stranded antibodies (anti-dsDNA) and anti-proteinase-3 (PR-3) antibodies (anti-neutrophil cytoplasmic autoantibodies cytoplasmic specific, cANCA) were assessed by the ELISA method. Fifty out of 86 patients had the systemic lesions. Pathological picture of the vascular loop in the nailfold capillary microscopy was found in 84 patients. In 19 patients the microvascular changes were advanced, in 35 moderate and in 30 mild. All patients with the articular manifestations had the pathological changes in the capillary microscopy. Patients with advanced changes in the capillary microscopy had the longer disease duration compared to patients with mild intensity of vasculitis. Serum concentration of sICAM-1 was significantly increased in RA and CTD patients compared to 30 controls (in both cases p<0.001). Moreover, RA and CTD patients with the systemic vasculitis showed significantly higher levels of sICAM-1 than those without vascular involvement (respectively p<0.001 and p<0.005). ANA were observed in significantly elevated concentration among RA and CTD patients with the systemic damage compare to patients without organ injury (respectively p<0.001 and p<0.05). Also cANCA level was twice more higher but only among CTD patients with the systemic damage (p<0.05). Serum concentration of sICAM-1 was elevated in studied patients with the presence of ANA antibodies (p<0.05). Significant correlation between ANA level and the disease duration, and hemoglobin concentration were observed. The concentration of cANCA correlated with rheumatoid factor and of dsDNA with patient age. We conclude that the systemic lesions in the course of RA and CTD are accompanied by the microvascular injury in nailfold capillary microscopy. Our data suggest that sICAM-1, ANA and cANCA serum levels may reflect the extent of the vascular involvement in RA and CTD patients.

Keywords: autoantibodies, sICAM-1, rheumatoid arthritis, connective tissue diseases, vasculitis, capillary microscopy.

49z603
Lactoferrin Stimulates Killing and Clearance of Bacteria but Does Not Prevent Mortality of Diabetic Mice.
Tadeusz Zagulski, Zofia Jarząbek, Alina Zagulska, Magda Jaszczak, Iwona E. Kochanowska and Michał Zimecki

Abstract. We have previously shown that bovine lactoferrin (BLF) given intravenously (i.v.) protected mice against a lethal dose of E. coli and strongly stimulated both the clearing and killing activities in liver, lungs, spleen and kidney. Since some studies indicated a reduction of the manifestation of experimental pancreatitis with lactoferrin, we decided to examine the protective activity of BLF against lethal E. coli infection in animals with alloxan (Alx)-induced diabetes. It appeared that 48 h diabetes substantially lowered the killing activity in all four organs as well as the clearing rate of E. coli from the circulation. BLF given i.v. o reduced this undesirable effect of diabetes. However, in 10- and 20-day diabetic animals, the diabetes alone stimulated the killing activity in the organs investigated, and upregulated the clearing rate of E. coli from the circulation. Lactoferrin (LF) significantly increased both the killing and the clearing activity in these long-term diabetic animals. In some cases the stimulating effect of BLF was very high, suggesting a concerted action of BLF and diabetes in that category of mice. Despite these beneficial effects of BLF and diabetes on the killing process in the investigated organs, the survival time of animals from all the diabetic groups ( 48 h, 10 and 20 days) was not prolonged by BLF. The protective properties of BLF did not depend on the blood glucose levels in the diabetic animals. BLF partly delayed the development of experimental Alx-induced diabetes, measured by the glucose level, but only if administered shortly after Alx injection. In conclusion, we demonstrated that the state of diabetes alone could increase killing of bacteria in the investigated organs and LF enhanced this process. However, LF had no protective effect against the mortality of diabetic mice infected with a lethal dose of E. coli.

Keywords: lactoferrin; protective activity; lethal infection; diabetes; E. coli; mice.

49z604
Investigation of Serum Cytokine Levels and Cytokine Production in Whole Blood Cultures of Paranoid Schizophrenic Patients.
Teresa Kamińska, Alicja Wysocka, Halina Marmurowska-Michałowska, Halina Dubas-Ślemp and Martyna Kandefer-Szerszeń

Abstract. There is some evidence that the pathophysiology of schizophrenia is related to changes in the innate and adaptive immune systems. In an attempt to define a potential immunological dysfunction in schizophrenia, we measured the serum levels of several cytokines in the sera of 24 patients with paranoid schizophrenia and investigated the cytokine production in whole blood assays after stimulation in vitro with virus (Newcastle disease), phytohemagglutinin (PHA) or bacterial lipopolysaccharide (LPS) and compared them with healthy, normal controls. A significant increase of IL-6, IL-8 and IFN-g levels, but a decreased IL-10 level were observed in the sera of patients with schizophrenia. No significant changes in the serum levels of IL-2, IL-4, IFN-a and TNF-a were detected in these patients. When cytokine production in vitro was examined, a significant defect in PHA-induced IL-2, IL-4 and IFN-a, and in virus-induced IFN-a production, but no significant alterations in LPS-induced IL-6, IL-10 and TNF-a production were observed. In summary, increased serum levels of some cytokines such as IL-6, IL-8 and IFN-g indicate an activation of the inflammatory response in schizophrenia, while the in vitro assay indicates significant changes in the Th1 (decreased production of IL-2 and IFN-g) and Th2 (decreased production of IL-4) cell system responses. The role of the defective IFN-a production in the regulation of the imbalance between Th1 and Th2 cell system responses is suggested.

Keywords: paranoid schizophrenia; cytokine production; IL-2; IL-4; IL-6; IL-8; IL-10; IFN-a; IFN-a; TNF-a.

49z605

Cellular Immunology

Influence of Dendritic Cells on the in Vitro Allogeneic Cytotoxic Reaction of Lymphoid Cells Derived from Normal or Listeria innocua-Infected BALB/c Mice
Teresa Gościcka, Maria Walencka, Marek Fol, Agnieszka Krupa and Jacek Szeliga

Abstract. The role of lymphoid dendritic cells (DCs) in the development of an allogeneic cytotoxic reaction in vitro was examined. The T+B and T cell subsets originating from the spleens or lymph nodes of normal and Listeria innocua-infected BALB/c mice were used as the effector cells. Their cytotoxicity to 51Cr-labeled C3H fibroblasts was determined after removal of DCs and replacing them again. Moreover, the influence of exogenous mrIL-12 on the potency of DCs in the allogeneic reaction developed in vitro was checked. It was found that the DC-deprived T+B or T subsets of splenocytes, regardless of their origin, exhibited 27-38% lower cytotoxicity than those accompanied by natural DCs. The cytotoxicity of these subsets from normal lymph nodes decreased by 22%, while the activity of bacteria-primed cells dropped by 38%. Replenishing effector cells with isolated DCs restored their cytotoxicity. Pulsation of normal DCs with IL-12 had no effect on the recovery of normal cell cytotoxicity. However, the IL-12-pulsed DCs were able to intensify the cytotoxicity of T+B subsets derived from the spleens or lymph nodes of L. innocua-infected mice. The results suggest that the alloantigen presentation by DCs to cytotoxic lymphocytes also takes place in the reaction developed in vitro, regardless of effector cell origin.

Keywords: dendritic cells; cytotoxicity; allogeneic reaction; Listeria innocua.
Effect of IFN-g on Expression of HLA in Bare-Lymphocyte Syndrome-Like Cell Line HAJ
Izabela Nowak, Bogusława Pochroń, Anita Kozłowska, Joanna Dubis and Piotr Kuśnierczyk

Abstract. We compared HLA antigen expression on new B-lymphoblastoid cell line (B-LCL) HAJ with that on B-LCLs expressing normal HLA levels as well as on B-LCLs derived from bare lymphocyte syndrome (BLS) patients and in vitro mutated B-LCLs of BLS-like phenotype. HAJ cells had no expression of HLA class II and low expression of class I antigens similarly to some of BLS B-LCLs, although HAJ cell line was derived from lymphocytes of HLA class I- and class II-normally expressing donor. HAJ cells displayed B lymphocyte markers, surface immunoglobulin and CD19. Culture of HAJ cells in the presence of interferon gamma resulted in HLA class I antigen upregulation, but did not restore class II expression. The cell line HAJ may prove useful for studies on factors influencing HLA class I cell surface expression.

Keywords: HLA expression defect, B-lymphoblastoid cell line, flow cytometry, interferon g.

49z607

Vol. 49, No. Supplement 1, 2001

Reviews

Regulation of Immunological Mucosal Tolerance
Georg Kraal (Department of Molecular Cell Biology, Vrije Universiteit, Amsterdam, The Netherlands) and Danielle A.W. Wolvers (Department of Immunology, UNILEVER Health Institute, Vlaardingen, The Netherlands)

Abstract. Mucosal tolerance is an immunological phenomenon specific to mucosal surfaces as found in the lungs and gastro-intestinal tract. It results in the suppression of immune responses to inhaled or ingested antigens and prevents the body from unwanted and unnecessary immunological responses to harmless molecules, such as grass-pollen or food constituents. This imposes the difficult task for the immune system of keeping a balance between reacting and non-reacting, and disturbances of this balance result in allergies and possibly autoimmunity, as well as opportunistic infections and even an escape from tumor surveillance. Understanding the mechanisms that underlie mucosal tolerance is, therefore, important from different viewpoints. Maintenance or (re)induction of mucosal tolerance to, e.g., food proteins, airborne allergens or autoantigens is desirable to prevent or cure allergies and autoimmune diseases. However, induction of mucosal tolerance is an unwanted phenomenon in mucosal vaccination and in the case of mucosal tumors.

Keywords: tolerance; mucosa; nasal; lymph node.

49s101

Liver Sinusoidal Endothelial Cells: a New Type of Organ-Resident Antigen-Presenting Cell
Andreas Limmer and Percy A. Knolle (Zentrum für Molekulare Biologie Heidelberg (ZMBH), Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany)

Abstract. The induction of peripheral immune tolerance in the liver is a well-known phenomenon that is operative in different situations such as tolerance to organ transplants and tolerance to oral antigens. The mechanisms leading to peripheral immune tolerance in the liver are still incompletely understood. While different cell populations of the liver have been implicated in and probably contribute in concert to the induction of hepatic immune tolerance, one hepatic cell type in particular seems to be suited for tolerance induction: liver sinusoidal endothelial cell (LSEC). LSEC are microvascular endothelial cells with a unique phenotype reminiscent of dendritic cells and a unique function as antigen-presenting cells for CD4+ T cells. The hepatic microenvironment, i.e. portal venous constituents and soluble mediators from sinusoidal cell populations, tightly control antigen presentation by LSEC to avoid immune-mediated damage. LSEC, in contrast to other endothelial cells, have the capacity to prime naive CD4+ T cells and induce cytokine release. Importantly, naive CD4+ T cells primed by antigen- presenting LSEC differentiate into regulatory T cells whereas T cells primed by bone marrow-derived professional antigen presenting cells differentiate into Th1 cells. Thus, LSEC represent a new type of organ resident “non-professional” antigen-presenting cell that appears to be involved in the local control of the immune response and the induction of immune tolerance in the liver.

Keywords: local immune response; antigen presentation; immune tolerance.

49s102
Two Receptor Theory in Innate Activation: Studies on the Receptors for Bacillus Culmet Guillen-Cell Wall Skeleton (BCG-CWS)
Tsukasa Seya, Misako Matsumoto, Shoutaro Tsuji, Midroi Nomura (Department of Immunology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Higashinari-ku, Osaka 537, Japan, The Organization for Pharmaceutical Safety and Research (OPSR), Tokyo 113, Japan), Nasim A. Begum (Department of Immunology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Higashinari-ku, Osaka 537, Japan), Ichiro Azuma (Hakodate National College of Technology, Tokura 14-1, Hakodate 042-8501, Japan), Akira Hayashi and Kumao Toyoshima (Department of Immunology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Higashinari-ku, Osaka 537, Japan)

Abstract. Activation of the innate immune system is a prerequisite for the maturation of dendritic cells (DC) and macrophages (Mo) followed by clonal expansion of the lymphocytes, targeting cells expressing „non-self” angitens. Microbes usually have a component competent to active DC/Mo for antigens presentation. This component has been colled adjuvant, but recently renamed pathogen-associated molecular pattern (PAMP) or modulin based on its molecular identification. Here, we propose the hypothesis that DC/Mo express two sorts of receptors for PAMP, whose signaling pathways lead to a sufficient antigen (Ag)-presenting state. In bacterial infection, a Toll-like receptor (TLR) and an uptake receptor participate in DC maturation and Mo activation. Likewise, with a number of viruses, two of the receptors with short consensus repeats (SCR), immunoglobulin-like domains or chemokine receptor-like motifs etc. induce functional modulation of DC/Mo. In immune therapy for cancer, primary activation of the innate system would be essential for tumor Ag-specific T cell augmentation. Cancer cells express tumor-associated Ag but barely co-express PAMP, which situation does not allow for the activation of innate immune responses. Supplementing tumor-associated Ag with PAMP may be an effective therapy for patients with cancer. Here, we discuss the possibility of an innate immune therapy for cancer with references to bacillus Culmet guillen cell-wall skeleton (GCG-CWS).

Keywords: modulin; pattern-recognition receptors; leucine-rich repeat (LRR); short consensus repeat (SCR); antigen-presenting cells (APC).

49s103
Regulation of Immune Responses by Natural Killer T Cells
Shayan Sharif and Terry L. Delovitch (Autoimmunity/Diabetes Group, The John P. Robarts Research Institute, and Departments of Microbiology and Immunology, and of Medicine, University of Western Ontario, London, Ontario N6G 2V4, Canada)

Abstract. Natural killer T (NKT) cells, which comprise a minor population of T cells in primary and secondary lymphoid organs, possess phenotypic characteristics of both NK and T cells. NKT cells respond to various external stimuli by an early burst of cytokines, including IL-4 and IFN-g. Thus, a key immunoregulatory role has been attributed to them. Autoimmune diseases, especially type I diabetes (TID), may be caused by dysregulation of the immune system, which leads to hyporesponsiveness of regulatory T helper 2 (Th2) cells and promotion of autoimmune Th1 cells. Furthermore, several lines of evidence exist to support the notion that an NKT cell deficiency in individuals at risk of TID may be causal to TID. As a result, targeting NKT cells using immunotherapeutic agents may prove beneficial in the prevention or recurrence of TID. Indeed, our data demonstrate that stimulation of NKT cells with a specific ligand prevents the onset and recurrence of TID in non-obese diabetic (NOD) mice.

Keywords: immunoregulation; autoimmunity; type I diabetes; alpha galactosylceramide.

49s104
The Social Life of NK Cells
Alfonso Martín-Fontecha (Department of Tumor Immunology, Scientific Institute San Raffaele, Via Olgettina 58, 20132 Milano. Italy) and Ennio Carbone (Microbiology and Tumor Biology Center, Karolinska Institutet, S-171-77 Stockholm Sweden; Cattedra di Immunologia, Dipartimento di Biologia e Patologia Cellulare e Molecolare, Universita di Napoli Federico II, 80131 Naples, Italy)

Abstract. Natural killer (NK) cells represent a distinct population of lymphocytes that was originally identified by its ability to kill transformed cell lines in vitro. It is now clear that these cells also play an important role in the innate immune response against a variety of pathogens, such as virus, bacteria and parasites. In the past few years, different protocols have been developed to activate NK cells ex vivo, allowing a detailed molecular analysis of the interaction of these cells with their cellular targets. NK activity is regulated by signals generated by both inhibitory and stimulatory receptors expressed by target cells. Indeed, recent results indicate that, while major histocompatibility complex class I molecules (MHC-I) expressed on target cells inhibit NK lytic activity by engaging surface inhibitory receptors, costimulatory molecules such as B7-1, B7-2 and CD40, are able to actively trigger NK activity. This review discusses the most recent findings on the role of costimulation on NK activation and forsees the possible consequences of the interaction between NK cells and dendritic cells (DC) on the development of an adaptive immune response.

Keywords: NK cells; costimulation; dendritic cells; B7-1; B7-2; CD40.

49s105

Local Tissue Complement Synthesis – Fine Tuning a Blunt Instrument
James E. Marsh, Wuding Zhou and Steven H. Sacks (Department of Nephrology and Transplantation, Floor 5, Thomas Guy House, Guy’s Hospital, King’s College, London, SE1 9RT, UK)

Abstract. Complement is important to host defense and the regulation of inflammation. The liver is overwhelmingly the major source of circulating complement. However, many other organs are capable of synthesizing some or all of the complement components in a regulated tissue-specific manner. There is increasing evidence that this locally generated complement is biologically active and exerts powerful effects within the local environment. We review the role of local complement synthesis within different organs and speculate on its implication for immune and metabolic functions.

Keywords: complement; kidney; brain; bone marrow; adipocyte; glomerulonephritis.

49s106
Role of Antigen-Presenting Cells in Innate Immune System
Toshiaki Ohteki and Shigeo Koyasu (Department of Microbiology and Immunology, Keio University School of Medicine, 35 Shinanomachi, shinjuku-ku, Tokyo 160-8582, Japan)

Abstract. Activation of antigen-presenting cells (APC) and natural killer (NK) cells initiates the production of various proinflammatory cytokines, including interleukin 12 (IL-12), interferon gamma (IFN-g) and nitric oxide (NO), which are important in the innate immune response for controlling infection by intracellular pathogens. In this review, we focus on these cytokines produced by APC and summarize the current understanding of how APC functions are regulated by cytokines in innate immunity.

Keywords: dendritic cells; macrophages; IL-12; IL-15; IFN-g; NO; gc; IL-2/15Rb.

49s107

Original papers

Effect of Cyclosporine A on the Nonspecific, Innate Antiviral Immunity of Mice
EWA ZACZYŃSKA and ZOFIA BŁACH-OLSZEWSKA (Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Weigla 12, 53-114 Wrocław, Poland)

Abstract. Different infections are the most common complication of immunosuppressive therapy. In this context, the effect of cyclosporine A (CsA) on the innate antiviral immunity of mice was studied. The presence of immunity was shown by infection of resident peritoneal cells (RPC) of BALB/c mice with herpes virus type 1 (HSV-1) and vesicular stomatitis virus (VSV). While the cells infected immediately after isolation were resistant to the viruses, the cells cultured for several days before infection lost immunity. The lack of activity to neutralize HSV-1 and VSV in the sera of the mice excluded a participation of specific antibodies in the resistance. To study the effect of CsA on innate immunity, BALB/ c mice were intraperitoneally (i. p.) injected with cyclosporine (20 or 100 mg/ mouse, twice a day) for three days. The other group of animals was injected in the same way with PBS only. Then the peritoneal cells were isolated and infected with VSV immediately after cell isolation. The kinetics of viral replication in the control and the CsA-treated groups was compared. While in the cells from the control group VSV did not multiply, in the cells from the CsA-treated mice the virus reached considerable titers. The cyclosporine effect on VSV replication was dose-dependent and statistically significant. We conclude that innate antiviral immunity was suppressed in the cyclosporine-treated mice and that this mechanism may be involved in the high susceptibility of patients to viral infections during immunosuppressive therapy.

Keywords: innate immunity; viral infections; cyclosporine.

49s108

Vol. 49, No. Supplement 2, 2001

Reviews

Is Tumor Expression of the Major Histocompatibility Complex Antigen Required for T Cell Immune Surveillance?
Yang Liu and Pan Zheng (Department of Pathology, Ohio State University Medical Center, Columbus, OH 43210, USA)

Abstract. Tumor expression of major histocompatibility complex antigen (MHC) class I and class II is not essential for the induction of memory T cells. However, induction of MHC class I-restricted effector cytotoxic T cells (CTL) appears dependent on MHC class I expression on tumors. Moreover, the effector function of tumor-specific CTL requires direct recognition of the tumor. In contrast, both the inductive and the effector phases of MHC class II-restricted T cells are independent of MHC class II expression on tumors.

Keywords: tumor immunity; major histocompatibility complex; immune surveillance; antigen-presentation; cross-priming

49s201
Chronic Hepatitis C Virus Infection and the Pathogenesis of Hepatocellular Carcinoma
Mark Feitelson (Department of Pathology, Anatomy and Cell Biology, Department of Microbiology and Immunology, Kimmel Cancer Center Thomas Jefferson University, Philadelphia, PA 19107, USA)

Abstract. There is a strong epidemiologic relationship between chronic hepatitis C virus (HCV) infection and the development of hepatocellular carcinoma (HCC), although the cellular and molecular mechanisms of tumor formation remain to be firmly established. Clearly, HCV is associated with the development of chronic hepatitis and cirrhosis, so that it may contribute to hepatocarcinogenesis as a consequence of its central role in the appearance and progression of necroinflammatory liver disease. There is also increasing evidence for a direct contribution of several HCV gene products to the development of the transformed phenotype, although none of the putative mechanisms involved in tumor formation have been strongly supported by in vivo evidence. Even if HCV is not shown to be a complete carcinogen, it may act as a cocarcinogen with underlying (serologically negative) hepatitis B virus (HBV) infection, in the context of alcoholic cirrhosis, and in patients with long term exposure to chemical hepatocarcinogens such as aflatoxin B1.

Keywords: hepatitis C virus; hepatocellular carcinoma; tumor pathogenesis

49s202
Immune Reaction to Breast Cancer: for Better or for Worse?
Helga M. Ögmundsdótti (Molecular and Cell Biology Research Laboratory, Icelandic Cancer Society, Reykjavik, Iceland)

Abstract. The infiltration of breast carcinomas with lymphoid cells has often been interpreted as an indication of an active immune response against the tumor and thus a favorable prognostic sign. Several studies have, however, cast doubt on this assumption. In situ breast carcinomas are more common than invasive cancers and it may be speculated that immune surveillance plays a role in preventing some localized cancers from becoming invasive. A secondary type of immune surveillance might be implicated in the long persistence of dormant breast carcinoma cells in the bone marrow. Breast cancer cells can carry tumor-associated antigens, particularly MUC1. These may elicit specific antibody responses but there is less evidence for a CTL response. There are indications that professional antigen presenting cells may be present and active at the edge breast tumours. Breast cancer cells may also interact directly with macrophages and NK cells. In terms of immune effector mechanisms in breast cancer, the communication with potential effector cells is likely to be often faulty because of altered expression of HLA class I molecules. Pleiotrophic cytokines are frequently present and could have a variety of effects ranging from growth inhibition to stimulated proliferation, loss of cell adhesion and activation of matrix degrading enzymes. Fas ligand is unlikely to play a role in the immune evasion of breast cancer. There is thus evidence for a variety of immune reactions to breast cancer. It is possible that they mediate some form surveillance, but growing, invasive tumors have escape routes and may even use cytokines to their advantage.

Keywords: breast cancer; immune surveillance; tumor antigens; cytokines; Fas-FasL; immune evasion

49s203

Original papers

Effect of Granulocyte-Macrophage Colony Stimulating Growth Factor on Interferon and Tumor Necrosis Factor Production in Whole Blood Cell Cultures of Patients with Acute Myelogenous Leukemia
Teresa Kamińska, Iwona Hus, Anna Dmoszyńska and Martyna Kandefer-Szerszeń

Abstract. The effect of recombinant human granulocyte-macrophage colony-stimulating growth factor (rHuGM-CSF) treatement on in vitro interferon (IFN) and tumor necrosis factor (TNF) production in peripheral blood cells of 46 patients with acute myelogenous leukemia (AML) was examined. GM-CSF significant enhanced virus-induced IFN-a production in blood cells (containing 70% of blasts) of 28 patients with M4-M5 AML according to the French-American-British (FAB) classification and also phytohemagglutinin (PHA)-induced IFN-g production in blood cells (containing 68% of blasts) of 18 patients with AML M0-M3 type. In control blood cells (25 healthy persons) GM-CSF enhanced PHA-induced IFN-g but did not influence IFN-a production. In the presence of GM-CSF, TNF-a titers induced with lipopolysaccharide were also higher in control blood cells but not in cells of patients with M0-M3 or M4-M5 type of AML. The significance of GM-CSF-enhanced IFN-a and IFN-g production in antimicrobial and antileukemic immune reactions which can develop during GM-CSF therapy is discussed.

Keywords: acute myelogenous leukemia; GM-CSF; interferon; tumor necrosis factor

49s204
Pre-Operative Levels of Serum Immunoglobulins, Circulating Immune Complexes and Complement Proteins in Patients with Different Types of Neoplasms
Hanna Gendek-Kubiak (1), Janina Grzegorczyk (2), Ewa G. Gendek (3), Marek L. Kowalski (2) and Jan Berner (4) – (1) Division of Cytophysiology, Department of Histology and Embryology, Medical University of Łódź, Narutowicza 60, 90-136 Łódź, Poland, (2) Department of Clinical Immunology and Allergy, Medical University of Łódź, Mazowiecka 11, 92-215 Łódź, Poland, (3) Institute of Technical Biochemistry, Technical University of Łódź, Stefanowskiego 4/10, 90-924 Łódź, Poland, (4) Department of Surgical Oncology, Medical University of Łódź, Paderewskiego 4, 93-509 Łódź, Poland

Abstract. In the sera of 30 neoplasm patients without metastases, the average IgG level was higher than in the control group (CG) (18.16 + 5.10 versus 12.62 + 2.14 g/l or 12.22 + 2.14 after excluding an out-lier). Average concentrations of CIC, IgM, C1i, C3c and C4 did not statistically differ between the groups. Dividing the patients’ group into breast or ovary cancer (BC), melanoma (M), digestive tract cancer (DT) and other neoplasms (ON) subgroups revealed that the IgG increase did not apply to the BC group. Relatively decreased CIC concentrations in the BC and DT group and an increased C1i in the DT group were found. Several diversities detected in the humoral immunity indices’ distributions and correlations suggest activation of different mechanisms depending on the neoplasm types.

Keywords: neoplasm(s); immunoglobulin(s); complement; immune complexes

49s205
Serum TNF-a Levels in the Neoplasm of Patients Qualified for Surgical Treatment
Hanna Gendek-Kubiak (1), Janina Grzegorczyk (2), Ewa G. Gendek (3), Marek L. Kowalski (2) and Jan Berner (4) – (1) Division of Cytophysiology, Department of Histology and Embryology, Medical University of Łódź, Narutowicza 60, 90-136 Łódź, Poland, (2) Department of Clinical Immunology and Allergy, Medical University of Łódź, Mazowiecka 11, 92-215 Łódź, Poland, (3) Institute of Technical Biochemistry, Technical University of Łódź, Stefanowskiego 4/10, 90-924 Łódź, Poland, (4) Department of Surgical Oncology, Medical University of Łódź, Paderewskiego 4, 93-509 Łódź, Poland

Abstract. The aim of the study was to analyze serum tumor necrosis factor alpha (TNF-a) levels in different neoplasm types qualified for surgical treatment and to evaluate their possible correlations with circulating immune complexes (CIC), IgG, IgM, and thecomplement (C) compounds: C1 inhibitor (C1i), C3c and C4 levels. Studies were performed in sera from 30 neoplasm patients before surgical treatment and in 10 persons from a control group (CG) with no malignancy. Serum TNF-a levels were measured with the Cytogen ELISA kit. Average TNF-a levels measured in neoplastic patient groups qualified for surgical treatment were not significantly different from the average TNF-a level in the CG group.

Keywords: neoplasm(s); TNF-a; serum; surgery

49s206
Characterization of NO and Cytokine Release by Transplantable Melanoma Cell Lines in Relationship to their Differentiation
Krystyna Kozłowska, Małgorzata Zarzeczna and Mirosława Cichorek (Department of Embryology, Medical University of Gdańsk, Dębinki 1, 80-211 Gdańsk, Poland)

Abstract. Changes in the secretory activity of two transplantable melanoma lines (differing in many biological features) as regards nitric oxide (NO) and interleukin 6 (IL-6), tumor necrosis factor a (TNF-a), oncostatin M (OSM) secretion were the subject of the present study. Obtained results show that a spontaneous alteration of the hamster’s native melanoma line in to an amelanotic one is accompanied by a global change of the secretory activity but there was not distinct correlation between the endogenous cytokine secretion and NO secretion by cells of both melanoma lines.

Keywords: NO; interleukin 6; tumor necrosis factor a; Oncostatin M; transplantable melanomas

49s207

Vol. 49, No. 1, 2001

DNA Tumor Vaccines

CD8+ T cell Suppressor Factors and the Control of Infection, Replication and Transcription of Human Immunodeficiency Virus

Phage Display as a Tool for Rapid Cloning of Allergenic Proteins

Inhibition of Angiogenesis in the Treatment of Tumors

Immunomodulatory Role of the Corticotropin Releasing Factor

Expression and Functional Significance of CTLA-4, a Negative Regulator of T Cell Activation

The Murine Ly49 Family: Form and Function

A Method for Directly Determining the Number of Dendritic Cells and for Evaluation of Their Function in Small Amounts of Human Peripheral Blood

Malformations of Angiogenesis in the Low Differentiated Human Carcinomas. Immunohistochemical Study

TNF-a, IL-6 and Their Soluble Receptor Serum Levels and Secretion by Neutrophils from Cancer Patients

Cytokine Production in Whole Blood Cell Cultures of Patients with B-Lineage Acute Lymphoblastic Leukemia. The Influence of Granulocyte-Macrophage Colony Stimulating Factor

Vol. 49, No. 2, 2001

CONTENTS

Review

Clinical Immunology

Immunochemistry

Vol. 49, No. 3, 2001

CONTENTS

Review

Cellular Immunology

Various

Vol. 49, No. 4, 2001

CONTENTS

Review

Clinical Immunology

Vol. 49, No. 5, 2001

CONTENTS

Review

Clinical Immunology

Vol. 49, No. 6, 2001

CONTENTS

Review

Clinical Immunology

Cellular Immunology

Vol. 49, No. Supplement 1, 2001

CONTENTS

Reviews

Original papers

Vol. 49, No. Supplement 2, 2001

CONTENTS

Reviews

Original papers