bluuebunny/crossref_metadata_2025 huggingface dataset:
A Study of Natural Cytotoxic (NC) Activity in the Metrial Glands of Rats and Mice
Authors: STEWART I. J. , PEEL S. , WEBSTER A. J.
Published: November 2003

Cell suspensions prepared from the metrial glands of rats killed on days 10, 13 and 20 of pregnancy and mice killed on day 12 of pregnancy were assessed for their ability to kill natural cytotoxic cell target Wehi 164 cells in a 51chromium‐release cytotoxicity assay. High cytotoxicity indices were found with metrial gland cell suspensions from rats killed on days 10 and 13 of pregnancy and from mice killed on day 12 of pregnancy. Cell suspensions from rats killed on day 20 of pregnancy, when there are relatively few of the granulated metrial gland cells which characterize the metrial gland, had little natural cytotoxic cell cytotoxic activity. Direct observation of mouse granulated metrial gland cells killing co‐cultured Wehi 164 cells were made using time‐lapse video recordings. The results indicate that granulated metrial gland cells are a type of natural cytotoxic cell.
Binding of Complement Proteins C1q and C4bp to Serum Amyloid P Component (SAP) in Solid Contra Liquid Phase
Authors: SØRENSEN I. JUUL , NIELSEN E. HOLM , ANDERSEN O. , DANIELSEN B. , SVEHAG S.‐E.
Published: November 2003

Serum amyloid P component (SAP), a member of the conserved pentraxin family of plasma proteins, binds calcium dependently to its ligands. The authors investigated SAPs interaction with the complement proteins C4b binding protein (C4bp) and C1q by ELISA, immunoelectrophoresis and electron microscopy. Binding of these proteins to SAP was demonstrated when SAP was immobilized using F(ab′)2 anti‐SAP, but not when SAP reacted with these proteins in liquid phase; thus the binding to human SAP was markedly phase state dependent. Presaturation of solid phase SAP with heparin, which binds SAP with high affinity, did not interfere with the subsequent binding of C4bp or C1q to SAP. In contrast, collagen I and IV showed partial competition with the binding of C1q to SAP. Using fresh serum, immobilized native SAP bound C4bp whereas binding of C1q/C1 could not be demonstrated. Altogether the results indicate that firm binding of C1q and C4bp to SAP requires that SAP is presented on a solid phase, that C1q and C4bp react with sites distinct from the heparin binding site, and that C1q and collagen I share binding sites on SAP. Immobilized native SAP, aggregated SAP and SAP‐heparan‐sulphate complexes induced no detectable complement activation.
Cytokine Production by Lymph Node Cells from Mice Infested with Ixodes ricinus Ticks and the Effect of Tick Salivary Gland Extracts on IL‐2 Production
Authors: GANAPAMO F. , RUTTI B. , BROSSARD M.
Published: November 2003

In BALB/c mice repeatedly infested with nymphal Ixodes ricinus ticks, lymphocytes from axillary and brachial lymph nodes which drain the tick attachment site produced significant levels of IL‐2, TNF‐α and GM‐CSF when stimulated in vitro with Con A or anti‐CD3 antibodies. Cytokine production by cells from lymph nodes of the opposite flank was equivalent to that of cells from uninfested mice. Nine days after the first infestation and IL‐2, GM‐CSF were produced primarily by the CD4+ T cells, while some other cell types contributed also to the TNF‐α production. In mice repeatedly infested, a gradual increase of lymph node cell production of IL‐2 was observed. The IL‐2 levels regularly increased from the first to the third infestation compared to TNF‐α levels which gradually decreased. The in vitro production of GM‐CSF was not affected by successive infestations. Spleen lymphocytes from naive mice produced higher levels of IL‐2 than lymphocytes from axillary and brachial lymph nodes. Both tick salivary gland extracts and D‐mannose inhibited IL‐2 production by these lymphocytes.
Cytokine Production in Hearts of Trypanosoma cruzi‐Infected CBA Mice: Do Cytokine Patterns in Chronic Stage Reflect the Establishment of Myocardial Pathology?
Authors: SUNNEMARK D. , ULFGREN A.‐K. , ÖRN A. , HARRIS R. A.
Published: November 2003

The authors analysed cytokine production in hearts of Trypanosoma cruzi‐infected CBA‐J mice by in situ immunocytochemical staining. Cellular infiltrates were recorded in hearts from both acute and chronic stages, but were not apparent in control hearts. In the acute heart, CD8 cells predominated, with associated production of IL‐4, IL‐6 and TNF‐α. Cytokine production was characterized by IL‐4, IL‐5, IL‐6 and TNF‐α in the chronic heart, and numbers of CD4 and CD8 cells were more equal. At this stage, calcified infarctions and associated fibrosis were apparent, mimicking chronic human Chagas' heart pathology. The authors consider the CBA mouse an appropriate model of chronic T. cruzi infection, and suggest that local cytokine production reflects establishment of heart pathology.
Altered Peptide Ligands of a Myasthenogenic Epitope as Modulators of Specific T‐Cell Responses
Authors: KIRSHNER S. L. , ZISMAN E. , FRIDKIN M. , SELA M. , MOZES E.
Published: November 2003

Myasthenia gravis (MG) is a T‐cell regulated autoimmune disease. A peptide representing a sequence of the human acetylcholine receptor α‐subunit (p195–212) was previously shown to stimulate proliferative responses of peripheral blood lymphocytes from MG patients and to be an immunodominant and myasthenogenic T‐cell epitope in SJL mice. The authors generated a panel of analogues of p195–212 that contain single amino acid substitutions. Three of the analogues (203PHE, 204GLY and 207ALA) triggered low to no proliferative responses of a p195–212‐specific T‐cell line designated TCSJL195–212. Two of these analogues were able to stimulate the line to produce interleukin‐2 (IL‐2) and IL‐4 (203PHE and 204GLY), whereas one analogue, 207ALA, did not stimulate the line to produce these cytokines. Binding assays revealed that the binding affinity of an altered peptide for a given major histocompatibility complex (MHC) molecule is not sufficient to determine whether it will be stimulatory or inhibitory to a native peptide‐specific T‐cell line. Two of the analogues, 204GLY and 207ALA, inhibited proliferative responses of cells of the TCSJL195–212 line when co‐cultured with p195–212 and syngeneic antigen presenting cells (APC). The two inhibitory analogues were also able to inhibit proliferative responses of the TCSJL195–212 line when APC were pre‐pulsed with p195–212, indicating that MHC blockade is not the only mechanism of action of these peptides. Moreover, both analogues inhibited the ability of p195–212 to prime lymph node cells for proliferative responses even when the analogues were administered in a soluble form. Thus the altered peptide ligands 207ALA and 204GLY can modulate T‐cell responses both in vitro and in vivo and may have therapeutic potential for the treatment of MG.
The Effect of Age on the Transendothelial Migration of Human T Lymphocytes
Authors: STOHLAWETZ P. , KOLUSSI T. , JAHANDIDEH‐KAZEMPOUR S. , KUDLACEK S. , GRANINGER W. , WILLVONSEDER R. , PIETSCHMANN P.
Published: November 2003

Since the migration of immunologically active cells into perivascular tissue is an important step in acute and chronic inflammation, the authors studied the possible influence of age on the transendothelial migration of T cells in an in vitro model. After incubation on collagen gels coated with human endothelial cells, three populations of T cells of elderly and young subjects were harvested: non‐adherent (NAD), bound (BND) and migrated (MIG) cells. The percentages of the BND and MIG fractions were similar in the young and elderly groups. Phenotypic analysis showed that in the starting population of the elderly subjects T‐cell subpopulations that have an increased capacity of transendothelial migration (e.g. CD4+ memory cells, CD8+/CD57+ cells) were enriched: nevertheless ageing was not associated with an increase in the transendothelial migration of T cells.
Bovine γ/δ TcR+ T Lymphocytes are Stimulated to Proliferate by Autologous Theileria annulata‐Infected Cells in the Presence of Interleukin‐2
Authors: Collins R. A. , Sopp P. , Gelder K. I. , Morrison W. I. , Howard C. J.
Published: November 2003

An in vitro model system has been developed in which freshly isolated resting WC1+γ/δTcR+ T cells proliferate in response to cells transformed by the protozoan parasite Theileria annulata, providing a strategy in which the basis of activation of naive γ/δ T cells can be investigated. Irradiated parasite‐transformed cells stimulate the proliferation, but not cytolytic activity, of autologous peripheral blood mononuclear cells (PBMC) from non‐immune animals. The proliferating cells are mainly WC1+γ/δ T cells. The majority of WC1+γ/δ T cells in freshly isolated PBMC express CD25 at a low level that increases when stimulated with T. annulata‐infected cells. Purified WC1+γ/δ T cells fail to proliferate when cultured with irradiated T. annulata‐infected cells and produce a small proliferative response to IL‐2, but proliferate strongly to irradiated or lightly fixed Theileria‐infected cells in combination with IL–2. The Theileria‐infected cells express cytokine transcripts encoding IL‐1α, IL‐1β, IL‐6 and IL‐10, but not IFNγ, IL‐2, IL‐4 and IL–7. Purified WC1+γ/δ T cells stimulated with T. annulata‐infected cells with or without IL‐2 fail to produce IL‐2 transcripts, but do produce those for TNFα. These experiments show that WC1+γ/δ T cells recognize a surface determinant on T. annulata‐infected cells, that together with a second signal, which can be provided by exogenous IL‐2, stimulates their proliferation.
Natural Polyreactive IgA and IgM Autoantibodies in Human Colostrum
Authors: VASSILEV T. L. , VELEVA K. V.
Published: November 2003

Secretory antibodies against bacteria and viruses in human colostrum and milk are known to be important protective factors for the breast‐fed infant. The authors have shown by enzyme immunoassay that colostrum contains IgA and IgM antibodies to a number of autoantigens: native DNA, actin, myosin, myoglobin, laminin, transferrin and thyroglobulin. These antibodies were polyspecific—those with anti‐DNA reactivity immunopurified on a DNA‐cellulose affinity column bound to a panel of self‐ and environmental antigens. The levels of natural autoantibodies in the immunoglobulin fraction of human colostrum were 3–10 times lower (when presented as antibody activity per μg of immunoglobulin) than in the immunoglobulin fraction of serum. The biological significance of the presence of B cells with autoantibody specificity in the mammary gland and of natural autoantibodies in colostrum and milk is not clear. It has been suggested that self‐reacting autoantibodies in serum play a major role in the selection of the pre‐immune B‐cell repertoire and in the maintenance of the immune homeostasis. The authors hypothesize that the natural autoantibodies in colostrum and milk may contribute to the selection process of physiological repertoire during the early postnatal period in breast‐fed infants. This could explain the lower frequency of allergic, inflammatory and autoimmune diseases and lymphomas which is seen in their later life when compared with that observed in children who have been formula‐fed after birth.
The Marginal Blood Pool of the Rat Contains not only Granulocytes, but also Lymphocytes, NK‐Cells and Monocytes: a Second Intravascular Compartment, its Cellular Composition, Adhesion Molecule Expression and Interaction with the Peripheral Blood Pool
Authors: KLONZ A. , WONIGEIT K. , PABST R. , WESTERMANN J.
Published: November 2003

To leave the blood, leucocytes marginate to the vessel wall. Granulocytes thereby form the so‐called marginal pool. It is unclear to what extent such a second intravascular compartment also exists for lymphocyte subsets, NK‐cells and monocytes. Samples of the peripheral blood and the marginal pool of the LEW rat were analysed by flow cytometry. In the marginal pool the percentage of granulocytes and monocytes was significantly higher compared to that of the peripheral blood, and the proportion of ‘naive’ T and B lymphocytes was decreased. The expression of LFA‐1 was higher on all leucocyte subsets of the marginal pool except the granulocytes, whereas no differences were seen for the expression of other adhesion molecules (α4‐integrins, ICAM‐1, CD2, L‐selectin, and CD44). In addition, splenectomy influenced the cellular composition of peripheral blood and marginal pool differently and, after injection of blood leucocytes, these cells were found in both compartments showing its characteristic cellular composition. Thus, not only granulocytes, but also B and T lymphocyte subsets, NK‐cells and monocytes form a second distinct intravascular compartment. This marginal pool probably influences the cellular composition of leucocyte subsets available for entry into the tissues.
Hierarchic T‐Cell Help to Non‐Linked B‐Cell Epitopes
Authors: BRONS N. H. C. , BLAICH A. , WIESMÜLLER K.‐H. , SCHNEIDER F. , JUNG G. , MULLER C. P.
Published: November 2003

The induction of antibodies against peptides requires the presence of a T helper cell epitope. In the absence of an added T‐cell epitope only 10% of the mice, or less depending on the strain, gave an antibody response to a series of peptides of the measles virus (MV) fusion (F) protein. After co‐immunization with a non‐covalently coupled T‐cell epitope more than 60% of the peptides became immunogenic. Considerable differences became apparent when BALB/c mice were immunized with peptides in the presence of different T‐cell epitopes. An immunodominant T‐cell epitope of the MV‐F protein was more efficient than a subdominant or a cryptic T‐cell epitope in providing help to a non‐linked B‐cell epitope. There is both a ranking order of the amount of help which B‐cell epitopes require and a ranking order for the help T‐cell epitopes are able to provide. The capability of a T‐cell epitope to provide help to a B‐cell epitope correlated with its own immunogenicity, i.e. the intensity of the antibody response to the peptide representing the T‐cell epitope. The data suggest that for each MHC class II allele there is an optimal T‐cell epitope which can provide help to a maximal number of B‐cell epitopes and that such a peptide can be identified by its ability to induce antibodies against itself. By using this strategy, the authors were able to induce antibodies which cross‐reacted with the MV.
Limited T Helper Cell Activity in C57BL/10 (B10) Mice with Inherited Low IgG Responsiveness
Authors: ŠÍROVÁ M. , ŘÍHA I. , ŘÍHOVÁ B.
Published: November 2003

Due to limitations in antigen processing, mice of the C57BL/10ScSn (B10) strain exhibit a low IgG production against a variety of T‐dependent antigens. To characterize the T‐cell functions, the authors studied antigen‐specific T‐cell proliferation and cytokine production in vitro. The response of B10 mice was compared with that of the high IgG producing strain A/J. A highly restricted proliferative response and almost no interleukin‐2 (IL‐2) and interleukin‐3 (IL‐3) production was detected in lymph node (LN) cells of B10 mice primed in vivo by protein antigens and subjected to a specific restimulation in vitro, whilst A/J cells responded by significant proliferation and cytokine production. The antigen‐specific T‐cell response of B10 mice could not be increased by lipopolysaccharide treatment in vivo or by in vitro cultivation with IL‐2. However, the T cells of B10 mice produced high levels of IL‐2 and IL‐4 when stimulated by phorbol myristate acetate (PMA) and Ca2+ ionophore, proving the existence of a functionally intact signal transduction pathway downstream of protein kinase C (PKC). The results suggest that the in vivo antigen priming does not effectively activate the T cells in B10 mice. The limited activation consequently leads to the low IgG response described in B10 mice.
CD8+ Cells are the Main Producers of IL10 and IFNγ after Superantigen Stimulation
Authors: HÖIDÉN I , MÖLLER G
Published: November 2003

Purified CD8+ T cells were recently shown to produce TH1 as well as TH2 types of cytokines upon restimulation, indicating an important role for these cells in regulation of immune responses. However, it is not known if the CD8+ cells would contribute to cytokine production in the presence of cytokine secreting CD4+ cells. In the present study the authors have investigated the proportion of cytokine‐producing CD4+ and CD8+ cells in the spleen after in vitro or in vivo stimulation. They found that stimulation of spleen cells with the superantigen Staphylococcal Enterotoxin B (SEB) in the presence of IL4 promoted production of IL10 and IFNγ predominately by CD8+ cells. In contrast, the production of IL4 was almost exclusively confined to the CD4+ subset. When priming with SEB in vivo before subsequent restimulation in vitro, a protocol previously shown to induce anergy, up to 80% of the IL10 and IFNγ positive cell expressed the CD8 marker. Taken together, these results emphasize the important role of cytokine‐producing CD8+ cells and indicate that CD4+ and CD8+ T cells may, in a given situation, produce distinct cytokines.
Expression and Regulation of Constitutive and Acute Phase Serum Amyloid A mRNAs in Hepatic and Non‐Hepatic Cell Lines
Authors: STEEL D. M. , DONOGHUE F. C. , O'NEILL R. M. , UHLAR C. M. , WHITEHEAD A. S.
Published: November 2003

‘Acute phase’ and ‘constitutive’ SAA (A‐SAA and C‐SAA, respectively) mRNA levels were measured in hepatic and non‐hepatic cell lines after treatment with monocyte conditioned medium (MoCM), with or without dexamethasone (Dex). A‐SAA mRNAs were detected in MoCM‐treated hepatoma cell lines (PLC/PRF/5, HuH7, HepG2, and Hep3B), a fibroblast cell line (MRC5), six epithelial cell lines (RT4/31, SW13, Hela Ohio, HCT‐8, CaCo2, and KB), and an endothelial cell line ECV304. In KB cells, Dex alone caused a dramatic increase in A‐SAA mRNA levels. C‐SAA was detected in all hepatic and non‐hepatic cell lines. Two differentially regulated size classes of C‐SAA mRNA were detected in the hepatoma cell lines. A‐SAA mRNA levels were measured in ECV304 cells treated with IL‐1β, IL‐6, TNFα and Dex, in various combinations, and revealed different profiles to those seen for hepatic cells. The extent of polyadenylation of A‐SAA mRNA in ECV304 and KB cells differed whereas the polyadenylation of C‐SAA mRNA remained constant. These data suggest that the parameters that determine the steady state mRNA levels and post‐transcriptional regulation of A‐SAA and C‐SAA mRNAs are different and are cell type specific.
Human Schistosomiasis: Modulation of In Vitro Granulomatous Hypersensitivity and Lymphocyte Proliferative Response by Macrophages Undergoing Differentiation
Authors: SILVA‐TEIXEIRA D. N. , DOUGHTY B. L. , GOES A. M.
Published: November 2003

The mechanisms associated with the modulation of immune response in the chronic phase of human schistosomiasis mansoni infection are complex and involve many cell types. In the present paper the authors demonstrate that antigenic stimulation of peripheral blood mononuclear cells (PBMC) from chronic‐intestinal schistosomiasis mansoni patients with polyacrylamide beads (PB) conjugated to Schistosoma mansoni soluble eggantigens (PB‐SEA) or adult worm antigen preparation (PB‐SWAP) were able to induce a statistically significant increase on the in vitro multinucleated giant cell (MGC) formation after the 15th day in culture. A correlation between an increase in the number of MGC and a decrease in in vitro granuloma formation index to PB‐SEA and PB‐SWAP was observed. Moreover, the authors demonstrated a down‐regulation of lymphocyte proliferative responses to S. mansoni antigens, during the differentiation pathway of monocytes towards MGC formation, due to a decrease in the antigen‐presenting capacity of these cells. These phenomena also correlate with a concomitant decrease in the expression of HLA‐DR and CD54 adhesion molecules on the surface of MGC. The results suggest that differentiation of monocytes to MGC may be one of the immunoregulatory mechanisms involved in the down‐regulation of the granuloma reaction against S. mansoni eggs.
Anti‐Interleukin‐6 Autoantibodies in Plasma are Associated with an Increased Frequency of Infections and Increased Mortality of Patients with Alcoholic Cirrhosis
Authors: HOMANN C. , HANSEN M. B. , GRAUDAL N. , HASSELQVIST P. , SVENSON M. , BENDTZEN K. , THOMSEN Å. C. , GARRED P.
Published: November 2003

Altered cytokine metabolism has been implicated in the pathogenesis of alcoholic liver disease. Recently, autoantibodies to cytokines have been proposed to act as modifiers of cytokine functions. In this study plasma levels of anti‐interleukin‐1α (IL‐1α autoantibodies and anti‐IL‐6 autoantibodies were determined by RIAs in 96 patients with alcoholic cirrhosis and in 16 healthy individuals. After 19 months (median) the prognostic significance of the cytokine autoantibodies was investigated using univariate analysis (Log‐rank test) and multivariate regression analysis (Cox model). The seroprevalences of anti‐IL‐1α autoantibodies and anti‐IL‐6 autoantibodies (42 and 18%, respectively) in the patients were not different from healthy individuals and did not relate to severity of liver disease. The presence of anti‐IL‐1α autoantibodies was of no prognostic significance. Independent of severity of liver disease, patients with anti‐IL‐6 autoantibodies in plasma had a higher risk of acquiring infections and higher risk of death (P < 0.02) compared to patients without anti‐IL‐6 autoantibodies. The authors concluded that anti‐IL‐6 autoantibodies are associated with increased mortality when present in the plasma of patients with alcoholic cirrhosis, which is probably secondary to recurrent infections, but not to underlying severity of liver disease.
A Role for L‐Selectin in Monocyte Activation by Jurkat Tumour Cells
Authors: Putz E. F. , MÄnnel D. N.
Published: November 2003

Monocytes and macrophages can distinguish between tumour cells and non‐malignant cells of the same cell type. The surface structures mediating tumour cell recognition and tumour cell‐induced cytokine production by monocytes or macrophages are still poorly characterized. The authors previously described N‐linked sialic acid‐containing carbohydrates associated with CD2 of the CD4+ tumour cell line Jurkat which induced tumour necrosis factor (TNF) secretion by human monocytes. In this report the authors demonstrate a role for monocytic L‐selectin in this process. Mannose‐6‐phosphate, an inhibitor of L‐selectin binding, blocked CD2‐induced TNF production whereas other ligands for L‐selectin (the monomeric monoclonal antibody to L‐selectin, DREG‐200; inositol hexaphosphoric acid; heparin) amplified CD2‐induced secretion of TNF. Polyvalent L‐selectin ligands as soluble monoclonal antibody (MoAb) DREG‐56, the cross‐linked MoAb DREG‐200, and the marine algal polysaccharide fucoidin induced TNF production without addition of CD2. Jurkat‐CD2 is associated with sialyl Lewis X‐related carbohydrates which differ in their expression or composition from that of the non‐stimulatory CD2 from non‐malignant T cells. Taken together the data presented in this report suggest that — at least in the case of Jurkat‐CD2 — L‐selectin is involved in monocyte activation by tumour typical carbohydrate structures.
Unresponsiveness of CD4+ T Cells from a Non‐Responder Strain to HgCl2 is Not Due to CD8+‐Mediated Immunosuppression: an Analysis of the Very Early Activation Antigen CD69
Authors: JIANG Y. , MÖLLER G.
Published: November 2003

Injections of HgCl2 lead to autoimmune manifestations in genetically predisposed rats and mice. In this study, the authors examined the responsiveness of T subsets from different mouse strains to HgCl2 by tracing their expression of the very early activation antigen CD69. The authors found increased expression of the CD69 antigen on CD4+ T cells from the responder A.SW and BALB/c mice, but not on CD4+ T cells from the non‐responder DBA/2 mice, indicating an activation of T helper cells in the responder strains. However, the CD69 antigen was induced on CD8+ T cells from all strains irrespective whether they were susceptible or resistant to mercury‐induced autoimmunity. Since CD8+ T cells have been described as mediating immunosuppression and as being responsible for the resistance to autoimmune induction by mercury, the authors tested whether CD8+ T cells inhibited the activation of CD4+ T cells by HgCl2 in the non‐responder strains. However, there was no evidence for a suppressive role of CD8+ T cells from the DBA/2 mice in the response to HgCl2. The findings indicate that T helper cells play a central role in the immunological effects of HgCl2 and unresponsiveness of T helper cells in the nonresponder strains is not due to CD8+‐mediated immunosuppression.
Analysis at the DNA Level of Human FcγRII Isoforms in Relation to the Polymorphic Binding of Murine IgG2b to Human B Cells
Authors: TAX W. J. M. , TAMBOER W. P. M. , MENSINK E. J. B. M. , KOENE R. A. P.
Published: November 2003

Three classes of human leucocyte Fcγ receptors (hFcγR) have been identified so far: hFcγRI, hFcγRII, and hFcγRIII. Previous studies have demonstrated that genetically determined differences between individuals exist both with respect to the binding of murine IgG1 (mIgG1) to hFcγ receptors, and with respect to the binding of murine IgG2b (mIgG2b). The polymorphism in binding of mIgG1 could be ascribed to hFcγRIIA, an isoform of hFcγRII. The authors have now investigated whether one of the isoforms of hFcγRII is also responsible for the polymorphism in binding of mIgG2b. In these studies the authors used EBV‐transformed human B cells that demonstrated either binding or no binding of mIgG2b in EA‐rosetting assays. mRNA obtained from these cells was amplified by reverse transcriptase and polymerase chain reaction (RT‐PCR). Hybridization experiments with the RT‐PCR products revealed that the hFcγRIIB but not the hFcγRIIA isoform was present in these cells. DNA sequencing further demonstrated that the nucleotide sequence of both the extracellular part and the cytoplasmic moiety of hFcγRIIB was identical for all individuals tested, regardless of their ability to bind mIgG2b. These findings indicate that the polymorphic binding of mIgG2b cannot be ascribed to one of the isoforms of hFcγRII. Since hFcγRI and hFcγRIII are not present on the cell surface of these cells, the authors conclude that an Fc receptor different from the known hFcγ receptors must be responsible for the polymorphic binding of mIgG2b. These data further expand the complexity of hFcγR.
VITAMIN A DEFICIENCY INCREASES INFLAMMATORY RESPONSES
Authors: Wiedermann U. , Chen X.‐J. , Enerbäck L. , Hanson L. Å. , Kahu H. , Dahlgren U. I.
Published: November 2003

The authors studied the influence of vitamin A deficiency on immediate and delayed type hypersensitivity as well as granulocyte‐mediated inflammatory reactions in vitamin A depleted and control rats. The number of circulating leucocytes was 43% higher in the vitamin A deficient than in the control animals. The leucocytosis was a result of a general increase of white blood cells and was not due to an increase in one particular type. The ratio between CD4+ and CD8+ T cells was unchanged. The vitamin A deficient rats had a four times higher T‐cell proliferative response and a two times higher interferon‐γ production in vitro than the control animals. In accordance, the DTH reaction was consistently higher in the vitamin A deficient rats. The granulocyte dependent inflammation, induced by olive oil injection, was also strongly enhanced in the vitamin A deficient rats compared with the controls. In addition, the spontaneous release of nitric oxide from the peritoneal phagocytes was five times higher in the vitamin A deficient animals. The number of peritoneal mast cells was about one and a half times higher in the vitamin A deficient than in the control animals. The density of IgE‐receptors on the mast cells, the IgE receptor occupancy and the histamine release from the mast cells did not differ between the groups, however. The vitamin A deficient immunized rats displayed a consistently stronger immediate skin reaction after intracutaneous antigen injection than the immunized control rats, despite lower IgE antibody levels. The skin reaction after intracutaneous injection of histamine was also significantly greater in the deficient animals. Despite the stronger reaction to antigen and histamine, the passive cutaneous anaphylaxis reaction was lower in the vitamin A deficient rats. In conclusion the study shows that vitamin A deficiency aggravates the clinical manifestations of inflammatory reactions. Thus, vitamin A deficiency might lead to a higher risk of acquiring irreversible tissue damage and disabling destruction.
GENOMIC ORGANIzATION AND PROMOTER ANALYSIS OF THE GENE ifngr2 ENCODING THE SECOND CHAIN OF THE MOUSE INTERFERON‐γ RECEPTOR
Authors: Ebensperger C. , RHEE S. , Muthukumaran G. , Lembo D. , Donnelly R. , Pestka S. , Dembic Z.
Published: November 2003

A clone containing the gene ifngr2 for the second chain (IFN‐γR2) of the mouse interferon γ receptor complex was isolated from a cosmid library made of 129/Sv mouse genomic DNA. Sequence analysis revealed that the second chain is encoded by 7 exons. The complete gene spans about 17 kb of the genomic DNA. In the 5′‐flanking region several transcription initiation sites between 27 and 136 nucleotides upstream from the translation initiation codon were mapped. This region has a high GC content, but no TATA or CAAT box. Potential binding sites were found for transcription factors Sp1, AP‐2, NF1, EGR and NFκB. Promoter activity was assayed with a series of constructs with firefly luciferase as a reporter gene, under the control of the promoter fragments of various lengths. This region showed promoter activity in transiently transfected Chinese hamster ovary cells.
Interleukin‐10 Suppresses the Development of Collagen Type II‐Induced Arthritis and Ameliorates Sustained Arthritis in Rats
Authors: PERSSON S. , MIKULOWSKA A. , NARULA S. , O'GARRA A. , HOLMDAHL R.
Published: November 2003

The collagen‐induced arthritis model in DA rats induced with homologous rat type II collagen was chosen to determine the therapeutic capacity and effects on autoimmunity by IL‐10. Systemic IL‐10 treatment (100 or 10 μg/day) with mini‐osmotic pumps during the periods of arthritis onset (days 12–20 after immunization) decreased the frequency of arthritis and delayed the onset and reduced the severity of arthritis in the few rats that eventually developed arthritis. Concomitantly, levels of autoantibodies to CII were reduced. To test the activity on established arthritis, IL‐10 was administered subcutaneously in the paws. This treatment reduced the swelling but did not block the arthritis process. The effective treatment required 100 μg of IL‐10 every 12th hour while 50 μg of IL‐10 had little effect, although a tendency of reduced paw swelling was observed. Surprisingly, therapeutic IL‐10 treatment led to higher serum levels of autoantibodies to CII. The highest doses of IL‐10 (100 μg) did not show any apparent toxic effects when given locally or systematically. Taken together, this study suggests that IL‐10 is a candidate for treatment of rheumatoid arthritis.
Monozygotic Twins Display Differences in Their Allospecific Cytotoxic T‐Cell Repertoire
Authors: ReisÆter A. V. , Brinchmann J. E. , Lundin K. E. A. , Thorsby E.
Published: November 2003

The antigen‐specific repertoire of peripheral blood T lymphocytes is generated by selection in the thymus followed by peripheral immunological events. While thymic selection is predominantly under genetic control, i.e. self‐HLA molecules + self‐peptides, confrontation with foreign antigens will lead to expansion of given T‐cells, thus modifying the T‐cell repertoire. To evaluate the relative importance of such modifying events the precursor frequencies (pf) of cytotoxic T lymphocytes (CTL) against given allogeneic HLA molecules were compared in six pairs of monozygotic twins. Using limiting dilution analyses, experiments with several different allogeneic stimulator/target cell donors were performed for each twin pair. In 12 out of 17 experiments, when the twins were tested against the same stimulator/target cell donor, the CTL pf differed and the 95% confidence intervals were non‐overlapping. This suggests that modifying effects by confrontation with foreign antigens play an important role in shaping the allospecific T‐cell repertoire.
MHC Class I Phenotype and Function of Human β2‐Microglobulin Transgenic Murine Lymphocytes
Authors: BJERAGER L. , PEDERSEN L. Ø. , BREGENHOLT S. , NISSEN M. H. , CLAESSON M. H.
Published: November 2003

Lymphoid cells from β2‐microglobulin (β2m) knockout mice transgenic for human (h) β2m (C57BL/10 mβ2m−/hβ2m+) were compared with normal mice for their binding to exogenously added hβ2m, binding to a H‐2Db peptide and for functional activity in a one‐way allogenic MLC. Based on data from cellular binding studies, Scatchard analyses and flow cytometry, it is concluded that exogenous hβ2m does not bind to hybrid MHC class I (MHC‐I) molecules composed of mouse heavy chain/hβ2m molecules expressed on lymphocytes of transgenic mice. Immunoprecipitation and SDS‐PAGE analysis of metabolically labelled normal C57BL/6 lymph node cells showed binding of exogenous hβ2m to MHC‐I, in particular, to the H‐2Db molecule through an exchange with endogenous mouse β2m. In contrast to normal H‐2Db molecules, hybrid H‐2Db expressed on the surface of transgenic lymphocytes binds radiolabelled peptide in the absence of exogenous added hβ2m suggesting that a stable fraction of hybrid H‐2Db molecules is empty or contain peptides with very low affinity. In a one‐way allogenic mixed lymphocyte culture, transgenic splenocytes were found to be far less stimulatory than normal splenocytes. In contrast, transgenic alloreactive cytotoxic T lymphocytes developed earlier in MLC than their non‐transgenic counterparts. These data indicate that the hybrid mouse heavy chain/hβ2m complex alters the alloantigenic repertoire and influences important aspects of T‐cell activation.
A Short History of Time and Space in Immune Discrimination
Authors: LANGMAN R. E. , COHN M.
Published: November 2003

The destructive effector functions of the immune system pose a problem that has aptly been described as ‘horror autotoxicus’. This problem demands a solution that offers an effective self–nonself discrimination mechanism. Unlike all other defence mechanisms, the immune system makes the self–nonself discrimination somatically, and not at the germline level. This discrimination requires a way of separating self from nonself. Two proposals to accomplish this are based on separation in time or in space. In this paper the authors show that separation in time remains the only viable solution. A generally accepted solution to the mechanism of the self–nonself discrimination is overdue as it strongly influences the way in which much of immune regulation is interpreted.
Effect of 12 Neutralizing Anti‐Cytokine Antibodies on In Vitro Activation of B‐Cells. Interleukin‐12 is Required by B1a but not B2 Cells
Authors: JONES B. M.
Published: November 2003

Normal human peripheral blood mononuclear cells, depleted of most monocytes and virtually all CD8‐positive cells, were stimulated in vitro with pokeweed mitogen plus Staphylococcus aureus Cowan I in the presence or absence of variousneutralizing anti‐cytokine antibodies. Numbers of CD5+ and CD5− immunoglobulin‐secreting cells were determined using the protein A haemolytic plaque assay after labelling B1a cells with anti‐CD5‐coated beads. Antibodiesagainst IL‐2, IL‐5 and IL‐10 had little or no effect on plaque‐forming cell (PFC) induction; anti‐IL‐6, ‐TNFα and ‐TGFβ enhanced PFC induction; anti‐IL‐1α, ‐IL‐1β, ‐IL‐4, ‐IFNγ and ‐IL‐13 suppressed PFC induction. B1a and B2 cells were equally affected by cytokine deprivation using these 11 neutralizing antibodies. In contrast, neutralizing anti‐IL‐12 suppressed induction of CD5+ but not CD5− PFC. Furthermore, recombinant IL‐12, if added during thefirst 48 h of culture, enhanced CD5+ PFC induction while marginally suppressing (IgG‐) or not affecting (IgA‐, IgM‐) induction of CD5− PFC. IL‐12 did not preferentially increase survival in culture of B1a cells norinduce expression of CD5 on B2‐cells. Further studies are required to determine whether manipulation of B1a and B2 subsets in vivo using IL‐12 could be achieved in clinical situations where imbalances in the two populations have beenobserved.
C5b‐8 Step Lysis of Swine Endothelial Cells by Human Complement and Functional Feature of Transfected CD59
Authors: MIYAGAWA S. , MIKATA S. , SHIRAKURA R. , MATSUDA H. , NAGASAWA S. , TERADO A. , HATANAKA M. , MATSUMOTO M. , SEYA T.
Published: November 2003

The authors established several swine endothelial cell (SEC) lines expressing human CD59 by transfection of cDNA, and assessed the function of the transfectant molecules in comparison with those of membrane cofactor protein (MCP) and decay‐accelerating factor (DAF) in an in vitro hyperacute rejection model of swine to human discordant xenograft. At the usual expression rate, DAF and MCP protected SEC from human complement mediated cell lysis, but CD59 did not block human complement attack on SEC. However, CD59 protects SEC from cell lysis when sufficiently expressed as in human umbilical vein (HUVEC). The authors examined why CD59 needed so many molecules to protect human complement‐mediated SEC lysis and found that SEC underwent lysis by human C5b‐8. The degree of C5b‐8 step lysis of SEC was approximately 70% of the total activity (C5b‐9). Additionally, CD59 protected human complement activation less efficiently at the C5b‐8 step than at the C9‐step. Therefore, to overcome human complement mediated SEC lysis, C8 activity must be inhibited by dense expression of CD59.
Inhibition by Rapamycin of P‐Glycoprotein 170‐Mediated Export from Normal Lymphocytes
Authors: Yacyshyn B. R. , Bowen‐Yacyshyn M. B. , Pilarski L. M.
Published: November 2003

P‐glycoprotein 170 encoded by the MDR‐1 gene mediates export of substrates including some immunosuppressive drugs. Rapamycin was compared to cyclosporine A for its ability to inhibit P‐glycoprotein on normal human peripheral blood mononuclear cells (PBMC). Rhodamine 123 dye efflux measures P‐glycoprotein activity and inhibition of P‐glycoprotein results in dye retention. Normal CD4+, CD8+ and B cells include a substantial subset with cyclosporine A‐sensitive rhodamine efflux. Rh123 dye efflux is also inhibited by rapamycin at comparable drug levels used in transplant models. CsA is approximately 100‐fold more effective on inhibition of PBMC P‐gp than is RAPA. P‐glycoprotein inhibition of ex vivo lymphocytes with three multi‐drug resistant T‐cell lines showed susceptibility of P‐glycoprotein to rapamycin dependent on the cell type. Compared to cyclosporine A, the reduced ability of rapamycin to inhibit P‐glycoprotein reflects a reduced avidity in its binding to P‐glycoprotein and perhaps increased access to the cell interior. The increased efficiency of RAPA as an immunosuppressive may in part be a result of its relatively low avidity for P‐glycoprotein. The authors speculate that interactions with P‐glycoprotein may partially modulate the immunosuppressive effects of rapamycin.
Recognition of B‐Cell Epitopes of the 65 kDa HSP in Behçet's Disease
Authors: DIRESKENELI H. , HASAN A. , SHINNICK T. , MIZUSHIMA Y. , VAN DER ZEE R. , FORTUNE F. , STANFORD M. R. , LEHNER T.
Published: November 2003

B‐cell epitopes of the mycobacterial 65 kDa heat shock protein (HSP) were mapped in sera from patients with Behçet's Disease (BD). A series of 47 overlapping synthetic peptides (15ers) derived from the sequence of the Mycobacterium tuberculosis 65 kDa HSP was used in ELISA. Significant increases in IgA and IgG antibody levels were observed with peptides 111–125, 154–172 and 311–326 in sera from BD, compared with those from controls. Homologous peptides derived from the sequence of the human mitochondrial 60 kDa HSP were then examined. Peptides 136–150 and 336–351 showed comparable results to the homologous mycobacterial peptides 111–125 and 311–326, respectively. The B‐cell epitopes defined in this investigation overlap with the T‐cell epitopes the authors have previously reported in BD. Inhibition studies are consistent with the view that antibodies to each of the three B‐cell epitope peptides represent a small proportion of the total B‐cell epitope repertoire elicited by the 65 or 60 kD HSP. Sequential antibody studies suggest that IgA and IgG antibody titres to one or all three peptides tested may increase during exacerbations of ocular disease. The functional role of these antibodies needs to be determined, but the peptides may be involved in the immunopathogenesis of BD as they can induce experimental uveitis in Lewis rats, which is a principal manifestation of BD.
Humoral Immune Responses Against Particulate Bacterial Antigens are Dependent on Marginal Metallophilic Macrophages in the Spleen
Authors: Buiting A. M. J. , De Rover Z. , Kraal G. , Van Rooijen N.
Published: November 2003

Lymphoid organs are populated by different macrophage subpopulations. In the spleen, four subpopulations can be characterized using differences in, for example, morphology, localization, cell markers and repopulation kinetics after liposome‐mediated depletion. The involvement of the different macrophage subpopulations in the immune response to particulate antigens was studied in vivo by intravenous injection of clodronate containing liposomes to eliminate the splenic macrophages, followed by immunization with trinitrophenylated Brucella abortus (TNP‐BA; thymus‐independent (TI)) or TNP‐Lactobacillus acidophilus (TNP‐LB; thymus‐dependent (TD)) at different time intervals. A strong decrease in the number of antibody‐secreting cells (ASC) against TNP‐LB was measured after elimination of all splenic macrophages. When TNP‐LB was injected after repopulation of the red pulp macrophages (completed after 2 weeks), the ASC response was still strongly reduced. Restoration of the capacity to mount an immune response correlated well with the reappearance of the marginal metallophilic macrophages. It is concluded that this particular subset of macrophages is involved in the immune response to these particulate bacterial TD antigens. In contrast, the response against the particulate bacterial TI antigen TNP‐BA was not impaired after depletion of splenic macrophages, although apparent changes in IgG isotypes were observed.
Differential Antigenic Stimulation Influences Cytokine Production Patterns in T Cells and CD4+ Subpopulations
Authors: EBTEKAR M. , KHANSARI N.
Published: November 2003

The regulatory mechanisms that govern the commitment of T cells to a Th1 or Th2 lineage in terms of cytokine production patterns have not yet been fully elucidated. The authors have endeavoured to study the role of the antigen in regulating the production of cytokines. To study this matter, a panel of antigens was chosen to include two random poly amino acids, PA1 (Poly (1‐PHe, L‐Glu) Poly‐dL‐Ala‐PolyL‐Lys), PA2 (Poly(Glu‐NaAla), and two purified protein derivatives PPD1 (H37Rv virulent) and PPD2 (H37Ra non‐virulent) obtained from WHO strains of Mycobacterium tuberculosis. After in vivo priming, murine spleen cells were prepared and three groups of cells (unfractionated, T cells, and CD4+ populations) were each separately stimulated in vitro with the original antigen Staphylococcal enterotoxin B (SEB) and phorbol myristate acetate (PMA). ELISA assays were subsequently performed on supernatants for IL‐4, IL‐5, IL‐2 and IFN‐g. The results indicate a different cytokine pattern for the various antigenic stimulations. The PPD1 induced IL‐5 production, while the PPD2 induced high levels of IFN‐γ. SEB was shown to exert a strong effect on the cytokine profile shifting it towards a Th1‐like profile. A comparison is made between the cytokine patterns in different cells. The role of antigens and superantigens in regulating cytokine production and determining the outcome of the pathological process in relation to other regulatory factors is discussed.
Suppression of Tumour‐Specific Cytotoxic T‐Cell Responses Against the Syngeneic BALB/c Plasmacytoma ADJ‐PC‐5 by Tumour‐Induced CD8+ Regulatory T Cells Via IFN‐γ
Authors: PAUELS H.‐G. , SPECHT C. , BECKER C. , KÖLSCH E.
Published: November 2003

The mechanisms of tolerance induction by tumour cells during early stages of tumourigenesis were analysed in a murine model system using the highly immunogenic BALB/c plasmacytoma ADJ‐PC‐5. Early stages of tumourigenesis were simulated in syngeneic BALB/c mice by repeated intraperitoneal injections with subimmunogenic doses of X‐irradiated ADJ‐PC‐5 tumour cells. This treatment causes a state of tumour‐specific tolerance in a high percentage of mice, involving a population of CD8+ peritoneal T cells which are able to suppress a protective tumour‐specific Tc response against this tumour. Using a primary mixed lymphocyte tumour cell culture (MLTC) as an in vitro system to study suppressive mechanisms of such regulatory T cells, the role of production or consumption of a number of cytokines was analysed. The data presented here demonstrate that inhibition of a protective Tc response against ADJ‐PC‐5 tumour cells is due to IFN‐γ production by suppressive T cells from tolerized mice, but not to IL‐2 consumption. In contrast to typical CD8+ Tc cells, ADJ‐PC‐5‐specific CD8+ Tc cells do not produce IFN‐γ and are furthermore suppressed by IFN‐γ. Thus, tumour‐induced suppressive T cells and tumour‐specific Tc cells seem to represent functionally and phenotypically different subsets of CD8+ T cells, possibly pointing towards a differential activation of type‐1 and type‐2 CD8+ T cells depending on the dose of tumour cells.
Heparin in Clinical Doses ‘Primes’ Granulocytes to Subsequent Activation as Measured by Myeloperoxidase Release
Authors: VIDEM V.
Published: November 2003

The author pre‐incubated isolated human granulocytes with increasing doses of heparin (0–100 IU/mL) before subsequent stimulation with formyl‐met‐leu‐phe. There was a dose‐dependant increase in granulocyte activation as measured by myeloperoxidase release, quantitated in enzyme‐immunoassay. This ‘priming’ effect of heparin was not due to activation of contaminating platelets or to endotoxin, and there was no reduction in granulocyte viability. Heparin‐induced ‘priming’ of granulocytes may be clinically relevant in the setting of cardiopulmonary bypass. The author describes a sensitive and specific double antibody enzyme‐immunoassay for myeloperoxidase from human granulocytes, with an inter‐assay coefficient of variation of 12.6%, an intra‐assay coefficient of variation of 4.3%, and a lower detection limit of 1.8 μg/l.
Anti‐Oxidants Inhibit the Enhancement of the Immune Response Caused by Oil Adjuvants
Authors: GIANNI P. DI , MINNUCCI F. S. , ALVES ROSA M. F. , VULCANO M. , ISTURIZ M. A.
Published: November 2003

Adjuvants are agents that can induce strong immunity to different antigens. They are thought to act mainly by stimulating macrophages, causing the release of cytokines, which in turn induce an inflammatory focus necessary for the adjuvant action. The authors found that catalase, ascorbic acid, N‐acetylcysteine and glutathione are able to inhibit the enhancing effect of incomplete Freund adjuvant (IFA) and polyoxyethylated castor oil upon the humoral immune response to sheep red blood cells (SRBC). None of the anti‐oxidants tested inhibited the basal immune response to the antigen. In addition, mice inoculated with different concentrations of hydrogen peroxide showed an enhanced response against SRBC, mimicking the effect observed with adjuvants. Delayed type hypersensitivity induced by SRBC in the presence of IFA was also inhibited by catalase. In conclusion, the report indicates that oxygen radicals are crucial molecules involved in the adjuvant effect observed in SRBC immunized mice.