bluuebunny/crossref_metadata_2025 huggingface dataset:
Gamma Interferon Induces the Appearance of a CD4‐, Major Histocompatibility Complex Class II+ Macrophage Subpopulation in the Rat Peritoneal Cavity
Authors: ERICSSON P. O. , HEDLUND G. , DOHLSTEN M. , CARLSSON R. , SJÖGREN H. O.
Published: June 2006

Peritoneal cells from gamma interferon (IFN‐γ)‐treated rats were phenotypically characterized by flow cytometry. Intraperitoneal inoculation of 3 ± 104 units of IFN‐γ induced, within 24 h, the appearance of a CD4‐, major histocompatibility complex (MHC) class II+ macrophage subpopulation not present in rats treated with phosphate‐buffered saline. IFN‐γ induced an increased expression of MHC class II I‐A molecules on both CD4‐ and CD4+ macrophages. Both cell types adhered to plastic and expressed high levels of the macrophage membrane molecules CD11b and OX41. Histological examination of sorted CD4‐ and CD4+ macrophages confirmed the macrophage morphology of both populations with less granula in the former. We conclude that the appearance of CD4‐ macrophages in the peritoneal cavity after inoculation with IFN‐γ most probably reflects a selective recruitment of these cells from blood or surrounding tissues. The function of these cells is still unknown, although strong expression of MHC class II I‐A indicates competence as antigen‐presenting cells.
Naloxone‐Reversible Monocyte Dysfunction in Patients with Chronic Fatigue Syndrome
Authors: PRIETO J. , SUBIRÁ M. L. , CASTILLA A. , SERRANO M.
Published: June 2006

We studied monocyte function in 35 consecutive patients with chronic fatigue syndrome (CFS) and 25 healthy controls. Eighty‐five per cent of the patients showed monocyte dysfunction characterized by marked reduction in the number of monocytes displaying immunoreactive cytoskeletal vimentin filaments, a low phagocytosis index, and a reduced expression of HLA‐DR antigens. These values increased dramatically after incubation of the patients' monocytes with the opioid antagonist naloxone. Other immunological abnormalities also noted in the patients were low lymphocyte blastogenesis and diminished numbers of monocytes displaying receptors for Fc of IgG (FcR) and C3b (CR1). These findings suggest that an increased opioid activity acting through a classical receptor mechanism is active on monocytes from a high proportion of patients with CFS and that this represents a novel example of immunomodulation by opioid peptides in human disease. We suggest that endogenous opioids are involved in the pathogenesis of the chronic fatigue syndrome.
Affinity Purification of IgG Subclasses and the Distribution of Thyroid Auto‐Antibody Reactivity in Hashimoto's Thyroiditis
Authors: WEETMAN A. P. , BLACK C. M. , COHEN S. B. , TOMLINSON R. , BANGA J. P. , REIMER C. B.
Published: June 2006

To delineate accurately the IgG subclass distribution of thyroid auto‐antibodies, sera from nine patients with Hashimoto's thyroiditis were fractionated into IgG subclasses by complete depletion of the other IgG subclasses on affinity columns. All IgG subclass fractions contained thyroglobulin and microsomal (or thyroid peroxidase) antibody activity, although when compared to the total serum concentrations of IgG subclasses. IgG4 antibodies were over‐represented. However, in contrast to recent studies, this particular subclass never predominated—IgG4 antibody levels being exceeded by those of the IgG1 and IgG2 subclasses: it seems likely that these differences relate to varying sensitivity for different subclasses in previously used assay methods. This pattern of subclass activity differed from that of tetanus toxoid antibodies, which were found in six subjects. There was no light chain restriction within any subclass, showing that the overproduction of IgG4 thyroid antibodies is not of monoclonal origin. The functional affinity of subclasses for both thyroid antigens varied between patients, but IgG2 subclass fractions showed the highest functional affinity in the majority of samples. We also found that IgG2 subclass thyroid antibodies were ineffective in eliciting antibody‐dependent cell‐mediated cytotoxicity, as distinct from the other three subclasses. Our results show that thyroid antibodies are less restricted in their IgG subclass distribution and patients are less heterogeneous than previously described. Moreover, IgG2 thyroid antibodies are quantitatively important and differ in relative functional affinity and effector function from IgG1 and IgG4 thyroid antibodies.
Human T Lymphocyte Clones: Influence of Culture Conditions and Optimization of Proliferative Assays
Authors: LUNDIN K. E. A. , BOSNES V. , GAUDERNACK G.
Published: June 2006

Many CD4+ human T lymphocyte clones (TLC) are found not to proliferate against appropriate stimulating cells, and many lose this capacity during culture. This may be due, not to a defect in the recognition of the antigen, but to an inability to produce sufficient amounts of interleukin 2 (IL‐2) for autocrine growth, since specific HLA‐restricted proliferative responses could be induced in ‘non‐proliferative’ clones by the addition of exogenous IL‐2 or phorbol myristate acetate (PMA). Of various factors tested during expansion procedures of the clones, the proliferative capacity could only be restored by changing the stimulatory cells from B lymphoblastoid cell lines (B‐LCL) to peripheral blood mononuclear cells (PBM). The cytotoxicity of the TLC was found to be independent of its proliferative capacity. After restoration of the proliferative capacity, a mouse B lymphoma cell line transfected with the appropriate HLA DQA and DQB genes was still not able to induce proliferation in the absence of exogenous IL‐2. We conclude that (1) ‘non‐proliferative’ TLC may recognize their targets, but fail to proliferate due to temporary lack of IL‐2 production, and (2) even ‘proliferative’ T cells may fail to respond to certain target cells carrying the specific antigen, such as a murine transfectant, in the absence of exogenous IL‐2.
Effect of Prolonged Monoclonal Antibody Administration on Cardiac Allograft Survival in the Rat
Authors: ROZA A. M. , KIMURA H. , MARKMANN J. , NAJI A.
Published: June 2006

After heterotopic cardiac transplantation in the rat, monoclonal antibodies (MoAb) specific for rat T‐cell subsets were administered until rejection. Across combined major histocompatibility complex (MHC) and non‐MHC differences (WF to Lew) and isolated non‐MHC differences (WF to Lew.1W) cardiac allografts were rapidly rejected in unmodified hosts (7.7 ± 1.0 days and 12.2 ± 0.8 days respectively). Across combined MHC and non‐MHC differences, administration of MoAb OX‐19 (pan T‐cell) on days ‐1,0, and 1 (where day 0 was the day of transplantation) and alternate days thereafter until rejection significantly prolonged allograft survival (28.5 ± 10.2 days, P<0.01). Administration of MoAb W3/25 (helper T cell) and MoAb OX‐39 (interleukin 2 (IL‐2) receptor) prolonged allograft survival (11.3 ± 2.6 days, P<0.05 and 13.3 ± 2.0 days, P < 0.01 respectively), whereas MoAb OX‐8 (cytotoxic/suppressor T cell) administration had no effect on allograft survival. In contrast, across non‐MHC differences (WF to Lew.1W) administration of MoAb OX‐8 markedly prolonged allograft survival (85, < 100 ± 3 days) whereas MoAb W3/25 administration had no effect. The effect of MoAb administration on lymphocyte subsets at rejection was assessed by flow cytometry. The relationship between depletion of targeted T‐cell subsets and graft survival was variable. Across both combined MHC and non‐MHC and isolated non‐MHC differences MoAb OX‐8 administration resulted in a marked reduction of OX‐8+ cells at rejection with no prolongation of graft survival in the former and indefinite graft survival in the latter. In contrast, OX‐19 administration resulted in prolonged graft survival but at rejection there were significant numbers of OX‐I9+ cells present. Administration of MoAb W3/25 failed to affect a significant reduction in W3/25+ cells, but allograft survival was nonetheless prolonged.
Regulation of IgG1 and IgE Synthesis by Interleukin 4 in Mouse B Cells
Authors: MOON H. B. , SEVERINSON E. , HEUSSER C. , JOHANSSON S. G. O. , MÖLLER G. , PERSSON U.
Published: June 2006

Mouse interleukin 4 (IL‐4) has been shown to act on B cells as an induction factor for Ig class switch. We studied the characteristics of IL‐4‐regulated Ig isotype production in lipopolysaccharide (LPS)‐stimulated splenic B‐cell cultures with emphasis on the comparison between the IgG1 and IgE responses. The results show that the kinetics for the appearance of IgG1 and IgE isotypes are similar, but that the dose of IL‐4 required for the induction of an IgE response is 3–10 times higher than that for an IgG1 response. No requirement for T cells was found for the induction of either isotype. Pre‐incubation of cells for 24 h with IL‐4 alone was sufficient 10 induce an IgG1 response when cells were recultured with LPS from days 1 to 6. However, the simultaneous presence of both IL‐4 and LPS for at least 24 h was required for a detectable IgE response. For an optimal IgE response. IL‐4 needed to be present for more than 72 h in LPS‐activated cultures. The possible reasons for the different regulation of IgG1 and IgE responses are discussed.
Immune Dysfunction in Primary Biliary Cirrhosis
Authors: CHIRICOLO M , LENZI M. , BIANCHI F. , FRANCESCHI C. , BARTOLINI G. , ORLANDI M. , TOMASI V. , LICASTRO F.
Published: June 2006

In a previous study we observed that after in vitro treatment with indomethacin, lymphocyte response to phytohaemagglutinin (PHA) in primary biliar) cirrhosis (PBC) patients was higher than that of controls. We know that indomethacin also inhibits prostanoid production, and thus in the present work we directly measured prostaglandin E2 (PGE2) and thromboxane B2 (TXB2) production by mononuclear cells and monocytes from 12PBC patients. 11 control subjects, and three control disease patients (alcoholic cirrhosis. AC). PHA‐stimulated enriched monocytes from PBC patients produced approximately threefold more PGE2 (after 48 h of culture) than did normal and AC monocytes (P<0.05). TXB2 production was similar in all groups studied. We also made cultures in which PBC‐purified lymphocytes proliferated better than PBC mononuclear cells (i.e. lymphocytes plus monocytes). Thus, a monocyte population producing PGE2 could be responsible, at least in part, for the hyporesponsiveness to PHA observed in PBC patients.
Selective Killing of CD4+ and CD8+ Cells with Immunotoxins Containing Saporin
Authors: BARBIERI L. , BOLOGNESI A. , DINOTA A. , LAPPI D. A. , SORIA M. , TAZZARI P. L. , STIRPE F.
Published: June 2006

Saporin, a ribosome‐inactivating protein from the seeds of Saponaria officinalis, was covalently linked to an anti‐CD4 monoclonal antibody. The resulting immunotoxin at 10‐9 M concentration was toxic to CD4+ lymphocytes without affecting other cells. Selective elimination of CD4+ and CD8+ cells was also obtained with murine monoclonal anti‐CD4 and anti‐CD8 antibodies and an immunotoxin consisting of saporin linked to an anti‐mouse IgG antibody.
Selection for Polymorphism in the Antigen Recognition Site of Major Histocompatibility Complex Class II Molecules
Authors: JONSSON A.‐K. , ANDERSSON L. , RASK L.
Published: June 2006

The genetic basis for the extensive polymorphism of major histocompatibility complex (MHC) class II molecules was investigated by statistical analysis. Nucleotide sequences of human DQA1, DQB1, DRB1, and DRB3 genes and murine Aα, Aβ, and Eβ genes were used. The results show that polymorphism is selected for in the antigen recognition site of class II molecules since replacement substitutions in this region were found to occur at a significantly higher frequency than expected in the absence of selection. In contrast, replacement substitutions are selected against in the remaining part of the first domain exon and in the second domain exon. Furthermore, comparing the sequence variability pattern among different class II α and β sequences, using a variability index for each residue, showed that, with few exceptions, highly polymorphic residues occur in the antigen recognition site. There was a strong and highly significant correlation in the variability pattern in the homologous DRB/Eβ sequences but not for DQB/Aβ or DQA/Aα sequences. This difference may be related to the fact that both α and β chains of DQ/A molecules are polymorphic, while only β chains of DR/E molecules vary.
Elevated IgG Antibody Levels to the Mycobacterial 65‐kDa Heat Shock Protein Are Characteristic of Patients with Rheumatoid Arthritis
Authors: TSOULFA G. , ROOK G. A. W. , BAHR G. M. , SATTAR M. A. , BEHBEHANI K. , YOUNG D. B. , MEHLERT A. , VAN‐EMBDEN J. D. A. , HAY F. C. , ISENBERG D. A. , LYDYARD P. M.
Published: June 2006

We have previously demonstrated raised levels of IgG and IgA antibody to the mycobacterial 65‐kDa heat shock protein (hsp) in the sera of patients with rheumatoid arthritis (RA). We have now attempted to determine whether this phenomenon is specific for RA, and whether it is seen only with the mycobacterial homologue of this particular hsp gene family. We therefore screened antibody levels to the mycobacterial and Escherichia coli hsp 65, and the mycobacterial, E. coli, and human hsp 70, in sera from RA, systemic lupus erythematosus (SLE), tuberculosis (TB), ankylosing spondylitis (AS), Crohn's disease, and control donors. RA sera show the greatest increase in IgA binding to the mycobacterial hsp65, but no increase in IgA binding to the E. coli homologue. Similarly, only RA and TB sera show increased IgG binding to the mycobacterial hsp65, and we have shown previously that the titre is greater in RA. In contrast, the use of mycobacterial and E. coli hsp70 preparations as control bacterial hsp gene products has shown that RA patients do not differ from TB or SLE patients in their antibody binding to these proteins. Moreover, neither IgA nor IgG antibody to the human hsp70 in RA sera were higher than in TB, and the IgA binding was not higher than in SLE. These findings suggest that elevated IgG antibody levels to the mycobacterial hsp65 shows some disease specificity, and further studies with the human homologue and at the T‐cell level are required.
Purification of Chicken C3 and a Structural and Functional Characterization
Authors: LAURSEN I. , KOCH C.
Published: June 2006

A major plasma protein from chicken, analogous to mammalian complement component C3, was purified by the removal of plasminogen, precipitation with polyethyleneglycol, and ion‐exchange chromatography. Purification was guided by a rabbit antiserum specific to chicken C3. The yield of native C3 was 27%, and purity and functional activity was assessed by SDS‐PAGE, immunoprecipitation techniques, and the ability of the purified C3 to restore the haemolytic activity of C3‐depleted chicken serum. Monoclonal antibodies were raised against purified chicken C3. These antibodies were characterized and used to prepare an immunosorbent column to deplete chicken plasma specifically of C3. Chicken C3 has a mol. wt of 185,000–195,000 and a two‐chain structure with an α chain (118,000) and β chain (68,000). Complement activation leads to changes in the electrophoretic mobility of chicken C3 and to a decrease in mol. wt to 144,000 corresponding to the release of a 15,000 C3a and a 34,000 C3d/C3dg fragment. Chicken C3 exists in multiple molecular forms with pI values of 6.4–6.6. A genetic polymorphism of chicken C3 based on electrophoretic mobility has not yet been detected after analysis of more than 500 individuals. The function of chicken C3 is dependent on a reactive thioester because treatment of purified chicken C3 with methylamine causes functional inactivation of C3.
Persistent Superphosphorylation of Leukosialin (CD43) in Activated T Cells and in Tumour Cell Lines
Authors: AXELSSON B. , PERLMANN P.
Published: June 2006

CD43 (leukosialin) is a highly sialylated, single‐chain molecule expressed on most human leucocytes. Regulatory signals appear to be transduced through the molecule as suggested by the ability of anti‐CD43 antibodies to induce aggregation and proliferation of T cells and to enhance B‐cell proliferation and natural killer cell activity. Activation of protein kinases is an essential event in signal transduction. We were therefore interested to study whether CD43 may function as a substrate for protein kinases during mitogenic activation of lymphocytes. We show that CD43 was rapidly superphosphorylated (within minutes) on serine residues following addition of phorbol ester (PMA) to peripheral blood lymphocytes. PMA treatment of the cells was not followed by rapid down‐regulation of CD43. Activation of the lymphocytes by concanavalin A or anti‐CD3 antibodies (OKT 3) also resulted in superphosphorylation of CD43. However, the phosphorylation was delayed as compared to that induced by PMA and was detected 3–4 h after the addition of the reagents. A plateau was reached after 24–48 h of Stimulation. Interestingly, the high level of phosphonrylation of CD43 was maintained in long‐term cultures of T cells activated by various means. Furthermore. CD43 was found to be constitutively superphosphorylated (on serine and tyrosine) in continuously growing cell lines of T, B, and non‐lymphoid origin. Taken together, the results suggest that CD43 has an important role during both early and late phases of T‐cell activation and that modulation of its biochemical properties by protein kinases may be associated with progression through the cell cycle and with cellular growth.
Functional and Molecular Characterization of a Monoclonal Antibody against the 165–186 Peptide of Human 1β
Authors: HERZBECK H. , BLUM B. , RÖNSPECK W. , FRENZEL B. , BRANDT E. , ULMER A. J. , FLAD H.‐D.
Published: June 2006

A synthetic peptide of human recombinant interleukin 1β (hrIL‐1β) 165–186, which exhibits biological activity in the human fibroblast assay, was used as an immunizing antigen to obtain a murine monoclonal antibody (MoAb) termed FIB 1 This MoAb, an IgG1, reacts specifically with hrIL‐1β. but not with hrIL‐1α, as measured in enzyme‐linked immunosorbent assays (ELISA). The MoAb FIB 1 detects the characteristic 17 kDa IL‐1 protein in Western blots. Binding to the antigen is specific, as deduced also from the close correlation of ELISA immunoreactivity with IL‐1 biological activity. The anti‐IL‐1β 165–186 Ab specifically neutralizes the biological activity of hrIL‐1β and native IL‐1, as measured by the IL‐1‐induced proliferation of murine thymocytes and human fibroblasts and the IL‐1‐dependent IL‐2 production by murine T cells (EL4‐6.1). Fifty per cent of hrIL‐1β activity (25 U/ml, or 0.25 ng/ml) has neutralized by less than 30μg/ml of MoAb. Furthermore, FIB 1 recognizes intracellular IL‐1 in lipopolysaccharide‐stimulated human peripheral blood mononuclear cells. The anti‐IL‐1β 165–186 Ab does not react with the shorter IL‐1β fragment 161–173 in solid‐phase ELISA, therefore the binding region seems to be localized in the amino acid sequence VALGLKEKN‐LYLS. A sandwich‐ELISA, using a polyclonal sheep anti‐IL‐1β 251–269 Ab as the capture antibody and anti‐1β‐165–186 MoAb as the detecting probe, allowed the determination of IL‐1β from crude culture supernatants.
A Monoclonal Antibody, H2, Defines a New Surface Antigen Expressed on Human Lymphocytes
Authors: GRUNEWALD J. , JANSON C. H. , TEHRANI M. J. , PORWIT A. , MATSUO Y. , MELLSTEDT H. , WIGZELL H.
Published: June 2006

We have previously described a monoclonal antibody (MoAb), H2, which recognized a tumour‐unique antigen on a human T‐cell chronic lymphatic leukaemia (T‐CLL, CD3,4+). However further characterization of H2 has revealed a reactivity with the majority of T lymphocytes and a minority of B lymphocytes, some malignant T cells and a few cell lines of leukaemia or of hematopoietic tumour origin. The molecular weight of the antigen (80,000) precipitated by the MoAb H2 from the cell lines NALM‐6 and Reh corresponded to that previously found. When PBL were stimulated with PHA, IL‐2, or Con A a reduced reactivity of H2 could be seen. The MoAb H2 was submitted to the Fourth International Conference on Human Leucocyte Differentiation Antigens, Vienna, 1989. H2 did not cluster in any of the 78 flusters of differentiation (CD 1–78) discussed at the conference, indicating its unique reactivity. This suggests that we have defined a new antigen on lymphocytes with a possible role along the resting proliferating axis.
Synovial Fluid from Rheumatoid Arthritis Patients Induces Polyclonal Antibody Formation in Vivo
Authors: ABEDI‐VALUGERDI M. , RIDDERSTAD A. , STRÖM H. , MÖLLER E.
Published: June 2006

Our previous studies demonstrated the presence of a T‐cell replacing factor in the synovial fluid (SF) of patients with rheumatoid arthritis (RA) and that RA‐SF can activate, selectively, the induction of IgG2b antibody secreting cells, in lipopolysaccharide (LPS)‐pretreated mouse spleen cell cultures. In the present study the effect of RA‐SF was tested in vivo in mice. Injection of the polyclonal activator LPS induced the production of IgM and IgG3 secreting cells in normal mice. However, the addition of RA‐SF led to a selective increase in the production of IgG2b with a peak response on day Sand IgG1 plaque‐forming cells (PFC) with a peak on day 7. Neither the IgG2b nor IgG1 responses were caused by specific immunity against heterologous proteins present in RA‐SF, as injection of in vitro inactive RA‐SF samples did not induce PFC. The effect on B cells of RA‐SF was further evaluated by injection of RA‐SF in combination with LPS to the Xid B‐cell deficient CBA/N mice, RA‐SF had identical effects in CBA/N as in normal mice. The biological implication of these findings is discussed. Our earlier results support the idea that B cells are endogeneously activated in RA patients, We have speculated that this activation is caused by the B‐cell differentiation factor which is present in SF. Therefore, we also tested whether RA‐SF could influence antibody‐forming cells in mice that spontaneously develop autoimmunity. We found that injection of RA‐SF alone, in the absence of any other activating substance, induced a very marked increase of IgG producing cells in (NZW × NZB) F1 hybrid mice. From a relatively high background level the RA‐SF could still induce an up to 100‐fold increase in the numbers of PFC in spleens of such mice.
Mitogen‐Induced Interleukin 2 and Gamma Interferon Production by CD4+ and CD8+ Cells of Patients with Inflammatory Arthritides. A Comparison between Cells from Synovial Fluid and Peripheral Blood
Authors: HOVDENES J. , GAUDERNACK G. , KVIEN T. K. , HOVDENES A. B. , EGELAND T.
Published: June 2006

The purpose of this study was to investigate interleukin 2 (IL‐2) and gamma interferon (IFN‐γ) production by purified CD4+ and CD8+ cells isolated from peripheral blood (PB) and synovial fluid (SF) of patients with rheumatoid arthritis (RA) and other inflammatory arthritides (non‐RA). CD4+ and CD8+ cells were selected positively by immunomagnetic separation. Supernatants of unstimulated CD4+ and CD8+ cells from both compartments did not contain any delectable IL‐2 or IFN‐γ, while supernatants of CD4+ and CDSI cells stimulated with phytohaemagglutinin and irradiated Raji cells mostly contained both cytokines. In vitro stimulated SF CD4+ cells gave supernatants with significantly less IL‐2 than supernatants from PBCD4+ cells, while in vitro stimulated SF CD4+ ‐cell supernatants contained significantly more IFN‐γ. SF CD4+ ‐cell supernatants contained significantly more IL‐2 than the parallel CD8+ supernatants, while there was no significant difference with regard to IFN‐γ content. The pattern of differences between SF ‐and PB‐derived T cells was the same for the two groups of patients, but the SF CD4+ cells from RA patients produced significantly less IL‐2 than the corresponding cells from the non‐RA group. The difference between SF and PB T cells with regard to lymphokine production is probably related to various degrees of in vivo pre‐activation. The results do not indicate a major T‐cell deficiency in relation to lymphokine production in RA.
Impaired Generation of Anti‐Viral Cytotoxicity against Lymphocytic Choriomeningitis and Vaccinia Virus in Mice Treated with CD4‐Specific Monoclonal Antibody
Authors: LEIST T. P. , KOHLER M. , ZINKERNAGEL R. M.
Published: June 2006

The role of CD4+ helper T cells in induction of anti‐viral cytotoxic T‐cell response was investigated by treating normal and thymectomized C57B1/6 mice with CD4‐speciflc monoclonal antibodies (MoAb). In CD4‐specific MoAb‐treated mice infected with Vaccinia or lymphocytic choriomeningitis virus (LCMV), cytotoxic T‐cell activity was 5–15 times lower than in normal controls when measured in a 51Cr release assay and computed as lytic units 6 and 8 days respectively after virus inoculation. This difference in the levels of effector T‐cell activities did not reflect slower kinetics of cytotoxic T‐cell induction in antibody‐treated versus control mice, since it was also obvious at 8 days after infection for Vaccinia virus and 10 and 12 days after inoculation with LCMV, CD4‐specific MoAb‐induced inhibition of cytotoxic T‐cell responses in vivo was seen up to 150 days after treatment in thymectomized mice. However, no significant suppressive effect of the same antibody treatment on T‐cell cytotoxicity could be observed in animals treated on day 3 or later after infection with Vaccinia virus. Injection of CD4‐depleted mice with recombinant interleukin 2 (rIL‐2) partially corrected the impaired virus‐specific cytotoxic T‐cell response, suggesting that IL‐2 supply may be limiting in mice licking T helper cells.
In Vitro Development of Bone‐Marrow‐Derived Macrophages Influence of Mouse Genotype on Response to Colony‐Stimulating Factors and Autocrine Interferon Induction
Authors: KRUSE A. , KIRCHNER H. , ZAWATZKY R. , DOMKE‐OPITZ I.
Published: June 2006

Responsiveness to granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) and macrophage CSF (M‐CSF) of bone marrow cells derived from different mouse strains was investigated. There were great variations in proliferation between different strains of inbred mice. Bone marrow cells from mouse strains with a high rate of proliferation in response to GM‐CSF also had a high proliferating capacity to M‐CSF. The response to either CSF did not correlate with a certain H‐2 haplotype. GM‐CSF induced consistently higher proliferation than M‐CSF. Proliferation in response to M‐CSF, but not to GM‐CSF, could be enhanced by the addition of antibodies against interferon (IFN). IFN is the only known inducer of (2′‐5′) oligoadenylate (oligo (A)) synthetase. This enzyme was induced in macrophages grown in the presence of M‐CSF, but not in GM‐CSF promoted cells. Enzyme induction was completely abrogated by simultaneous treatment with anti‐IFNα/β. Infection of macrophages with herpes simplex virus type 1 (HSV) and vesicular stomatitis virus (VSV) revealed that GM‐CSF‐promoted cells were highly susceptible to lytic infection by these viruses. In contrast, virus titres in M‐CSF‐cultured cells were 100‐fold lower. We conclude that, contrary to M‐CSF, GM‐CSF does not induce autocrine IFN during haematopoiesis. As judged from data with BALB/c mice, the sensitivity to the anti‐proliferative effect of the autocrine IFN may be a factor which influences M‐CSF‐promoted proliferation.
Rheumatoid Synovial Fluid Reconstitutes the B‐Cell Defect in CBA/N Mice
Authors: RIDDERSTAD A. , ABEDI‐VALUGERDI M. , STRÖM H. , MÖLLER G. , MÖLLER E.
Published: June 2006

Synovial fluid from patients with rheumatoid arthritis (RA‐SF) contains a biological activity which can replace T cells for activation of antibody secretion in human blood lymphoid cells and which can also induce the selective differentiation of IgG2b‐secreting cells in lipopolysaccharide (LPS)‐pre‐activated mouse spleen cells. The B‐cell activity of this factor was studied in CBA/N mice which have an X‐linked B‐cell immunodeficiency which manifests itself as a defective humoral response to certain thymus‐independent antigens (TI‐2). RA‐SF has now been shown to reconstitute partly the B‐cell deficiency in CBA/N splenic B cells in vitro Addition of RA‐SF to LPS‐pretreated cell cultures results in IgG2b secretion in CBA/N spleen cells as well. In contrast to cells from normal CBA mice, cells from CBA/N mice cannot respond to interleukin 4 (IL‐4) after addition of LPS with production of IgG1 antibodies in vitro. However, the addition of RA‐SF completely restores a normal IL‐4‐induced IgG1 response. No other biologically active factors have been shown to allow the production of IgG antibody producing cells in CBA/N splenic B cells. It is postulated that the xid immunodeficiency could be the result of a deficient production of a biological activity which is abundant in RA‐SF.
Synthesis of Complement by Alveolar Macrophages from Patients with Sarcoidosis
Authors: PETTERSEN H. B. , JOHNSON E. , MOLLNES T. E. , GARRED P. , HETLAND G. , OSEN S. S.
Published: June 2006

Sarcoidosis is a granulomatous disorder of unknown aetiology. Alveolar macrophages (AM) in sarcoidosis release a variety of mediators important to the pathogenesis of the disease. Complement is essential for the inflammatory response and we investigated whether there were any major defects in the potential for sacroidosis AM to synthesize complement in vitro. AM from 11 patients with active sarcoidosis and three healthy controls were cultured under serumfree conditions, There was a significant binding of polyclonal (anti‐C5, ‐C6, ‐C7, ‐C8) and monoclonal anti‐complement antibodies (anti‐C3c and anti‐C9 neoepitope (aE11)) to agarose beads incubated with unstimulated AM for 24, 48, or 72 h, A significant and inhibitable production of soluble C3c, C5, C9, and S‐protein was found in the harvested medium as detected by enzyme immunoassays. Activated C3 and C9 were also detected on neoepitope expression. Presence of co‐cultured agarose beads reduced the amount of soluble S‐protein due to deposition on the agarose. We argue that the C9 neoepitope is an integral part of the terminal complement complex (TCC), both in the fluid and solid phase when bound to the agarose. In the fluid phase. SC5b‐9 was generated, whereas the agarose‐bound S‐protein is assumed not to be associated with TCC on the beads. The results demonstrate for the first time that AM from sarcoidosis patients synthesize the functional alternative and terminal of complement.
Increased Amounts of C4‐Containing Immune Complexes and Inefficient Activation of C3 and the Terminal Complement Pathway in a Patient with Homozygous C2 Deficiency and Systemic Lupus Erythematosus
Authors: GARRED P. , MOLLNES T. E. , THORSTEINSSON L. , ERLENDSSON K. , STEINSSON K.
Published: June 2006

Plasma and serum samples from a patient with homozygous C2 deficiency and severe systemic lupus erythematosus who responded with full clinical remission after plasma infusions were examined for immune complexes (IC), C3 activation products, and the terminal complement complex (TCC). Plasma contained large amounts of C4‐containing IC but no C3‐containing IC or complement activation products. Classical pathway activation in vitro did not lead to C3 activation or TCC formation as seen in normal serum, but a very efficient binding of C1q and C4 was found. No disturbances in alternative pathway activation were observed. The results indicate an impaired formation of C3‐containing IC and an inefficient clearance of C4‐containing IC, supporting the idea of a causal relationship between the dysfunctional classical pathway, pathophysiology, and clinical manifestations in this patient.
Effects of Heat Shock on Cytolysis Mediated by NK Cells, LAK Cells, Activated Monocytes and TNFs α and β
Authors: JÄÄTTELÄ M.
Published: June 2006

We have recently shown that a heat treatment of a murine target cell line, WEHI 164, induces resistance to lysis mediated by tumour necrosis factors alpha (TNF‐α) and beta (TNF‐β). In the present study the effect of the heat shock of target cells on cytotoxicity mediated by natural killer cells (NK cells), lymphocyte‐activated killer cells (LAK cells), activated monocytes, TNF‐α, and TNF‐β was investigated, First, WEHI 164 cell line and six human cell lines (ME 180, K 562, U 937, HeLa, MCF 7, and SK‐OV 3) were screened for their sensitivity to different forms of lysis, and then sensitive cell lines were heat‐treated. Pretreatment of target cells at 42° C for 45‐60 min also rendered human target cell lines more resistant to lysis by rTNFs, and the acquired resistance was accompanied by an increased resistance to activated monocytes, but not to NK cells or LAK cells. Thus, the heat‐induced resistance mechanisms capable of protecting target cells from lysis by rTNFs and by activated monocytes do not elicit resistance to lysis by NK cells and LAK cells, supporting the hypothesis that mediators other than TNFs are involved in NK cell‐and LAK cellmediated killing.
CD5 mRNA Expression and Auto‐Antibody Production in Early Human B Cells Immortalized by EBV
Authors: PAAVONEN T. , QUARTEY‐PAPAFIO R. , DELVES P. J. , MACKENZIE L. , LUND T. , YOUINOU P. , LYDYARD P. M.
Published: June 2006

A number of studies have suggested that lymphocytes producing polyreactive antibodies belong to the CD5+ B‐cell subset. In this study we have examined CD5 at the cell surface and mRNA levels in EBV‐driven cord blood and fetal liver clones previously characterized in terms of their antibody specificities. We show that EBV‐immortalized cells can express surface CD5, and that some of the clones not expressing surface CD5 express it at the mRNA level. The complete absence of CD5 mRNA in some polyreactive clones is consistent with the proposition that the production of auto‐antibodies and multispecific antibodies is not restricted to the CD5+ B‐cell subset.
Presence of Human 65 kD Heat Shock Protein (hsp) in Inflamed Joints and Subcutaneous Nodules of RA Patients
Authors: KARLSSON‐PARRA A. , SÖDERSTRÖM K. , FERM M. , IVANYI J. , KIESSLING R. , KLARESKOG L.
Published: June 2006

Monoclonal antibodies to the human homologue of the bacterial 65 kD heat shock protein (hsp) were used to investigate the tissue distribution of endogenous hsp 65 in normal versus rheumatoid synovial tissue, in subcutaneous nodules of patients with rheumatoid arthritis (RA) and in several instances of non‐rheumatoid inflammation. A strong reactivity of the anti‐hsp antibody was found in the cartilage‐pannus junction in rheumatoid joints and in rheumatoid nodules, but not in normal joints or in normal or inflamed kidney or liver (irreversible graft rejection, chronic glomerulonephritis or primary biliary cirrhosis). The findings provide a new hypothetical explanation for a role of T cells reactive with the 65 kD hsp in the generation of both articular and extra‐articular lesions in chronic rheumatoid arthritis.
Purification and Characterization of Hamster Serum Amyloid A Protein (SAA) by Cholesteryl Hemisuccinate Affinity Chromatography
Authors: NIEWOLD TH. A. , TOOTEN P.C.J.
Published: June 2006

High‐density lipoprotein (HDL) apolipoprotein was separated from hamster serum by cholesteryl hemisuccinate atlinity chromatography (CHAC) in comparison with the density‐gradient ultracentrifugation (DGUC), The apolipoprotein recovery from scrum by CHAC was 70% and by DGUC 80%, This disadvantage is compensated for by the ease of purification by CHAC, a method particularly suited for the processing of large amounts of serum.

From the acute‐phase HDL CHAC fraction, apo SAA was isolated by gel filtration. Using isoelectrofocusing, two‐dimensional gel electrophoresis, und titration curve, four isotypes of hamster apo SAA were identified and characterized In the acute‐phase serum, one of the isotypes was predominant (apo SAA1), In serum of amyloidotic animals, the relative contribution of apo SAAl was considerably lower, suggesting selective removal of the latter during amyloidogencsis and possibly its deposition in hamster AA amyloid. Furthermore, the affinity chromatography method was modified with gradient elution of affinity‐bound material. By this method HDL apolipoprotein was separated into three subclasses. Apo SAA was shown to associate with two different subclasses. In acute‐phase serum most of the apo SAA1 was found in the subclass with the lowest affinity for the cholesteryl beads, whereas thc latter was depressed in amyloidotic serum, suggesting that the amyloidogenicity ofa particular apo SAA isotype is determined by its cholesteryl‐binding properties.
The Vδ Gene Usage by Freshly Isolated T Lymphocytes from Synovial Fluids in Rheumatoid Synovitis: a Preliminary Report
Authors: SIOUD M. , KJELDSEN‐KRAGH J. , QUAYLE A. , KALVENES C. , WAALEN K. , FØRRE Ø. , NATVIG J. B.
Published: June 2006

Taking advantage ofthe polymerase chain reaction we have studied the usage of variable delta‐(Vδ)) region genes in freshly isolated synovial fluid T cells from patients with rheumatoid synovitis. Amplified mRNA from one patient with rheumatoid arthritis (RA) was cloned into an Smaleleaved pUC19 vector and colonies were screened with probes for three of the known human variable δ‐gene families (Vδ1. Vδ2. Vδ3). Of 10 clones, seven used Vδl. two Vδ2 and one Vδ3. This pattern of distribution is different from that of normal peripheral blood, where approximately 60% of Tγδ) cells are reported to use the Vδ2 gene. Furthermore, Northern blot hybridization analyses of mononuclear cells from two additional synovial fluids derived from another patient with RA and one with juvenile rheumatoid arthritis (JRA) also showed significant hybridization only with Vδ I. In summary, these preliminary results suggest a usage of Vδ gene families in Tγδ lymphocytes in synovial fluid of rheumatoid patients different to that found in normal peripheral blood.
In Vivo and In Vitro Antibacterial Activity of Conglutinin, a Mammalian Plasma Lectin
Authors: FRIIS‐CHRISTIANSEN P. , THIEL S. , SVEHAG S.‐E. , DESSAU R. , SVENDSEN P. , ANDERSEN O. , LAURSEN S. B. , JENSENIUS J. CHR.
Published: June 2006

Conglutinin is a mammalian C‐lype lectin which agglutinates iC3b‐coated erylhrocytes, Ingram [13] found that euglobulin from bovine serum may confer partial protection against experimental infections in mice. We now present evidence that the protective activity in euglobulin against infections of BALB/c mice with Salmonella typhimurium is mediated by conglutinin. Conglutinin also demonstrated antibacterial activity against E. coli and S. typhimurium in vitro. The expression of this activity required the presence of heat‐labile serum factors and peritoneal exudate or spleen cells, but not antibodies to the bacteria. Antibacterial activity was also demonstrated when the bacteria were prelreated with serum at 37°C before incubation with conglutinin and cells. The activity of conglutinin was not observed when factor I‐deficient or EDTA‐treated serum was used instead of normal serum. The active peritoneal exudate or spleen cells showed adherence to plastic.
Analysis and Characterization of the Interleukin 2 Receptor α and β Chain Expression inCD4−CD8− Cells
Authors: GUTIÉRREZ‐RAMOS J. C. , PEZZI L. , LEONARDO E. , MARTIN C. , MARTINEZ‐A C.
Published: June 2006

The differential effects that the binding of interleukin 2(IL‐2)to its β or αβ receptors might induce in two different CD4− CD8− T‐cell lines were analysed. While LDl. T3b, a double‐negative T cell derived from MRL/Ipr mice, constitutively expressed high levels of the IL‐2R β chain, YAC‐1, a Moloney sarcoma virus‐transformed CD4− CD8−T cell, expressed (as an activated T cell) the β and α chains. The presence of IL‐2 in the culture medium was lethal for LDl. T3b cells, while it had no effect on the growth of YAC‐1 cells. IL‐2 increased the exprcssion of the β chain and, to a lesser extent, of the α chain in YAC‐1 cells. In addition, other markers such as CD4 and CD5 were induced by IL‐2 in this celt line.
The Neuropeptide Substance P Stimulates Production of Interleukin 1 in Human Blood Monocytes: Activated Cells are Preferentially Influenced by the Neuropeptide
Authors: LAURENZI M. A. , PERSSON M. A. A. , DALSGAARD C.‐J. , HAEGERSTRAND A.
Published: June 2006

Substance P (SP) has recently been reported to induce interleukin 1 (IL‐1) production by human monocytes. This was confirmed in our experiments with human monocytes cultivated in the presence of SP or SP together with lipopolysaccharide (LPS). In addition, a wide variability of cell response to the neuropeptide was noticed. Three out of twelve cell cultures were directly stimulated by SP to release IL‐1, while four additional cultures needed prestimulation with suboptimal doses of LPS, and no effect was seen in the five remaining experiments. The data may suggest that preferentially activated monocytes respond to SP. The production of IL‐1 by SPstimulated monocytes is of great interest considering the broad spectrum of activity of IL‐1 and the increasing evidence of sensory neuron involvement in acute and chronic inflammatory responses.
RT7‐Defined Alloantigens in Rats are Part of the Leucocyte Common Antigen Family
Authors: KAMPINGA J. , KROESE F. G. M. , POL G. H. , OPSTELTEN D. , SEIJEN H. G. , BOOT J. H. A. , ROSER B. , NIEUWENHUIS P. , ASPINALL R.
Published: June 2006

Haemopoietic cells carry a variety of cell‐surface molecules, some of which are known to have allotypic variation. In rats, the RT7 alloantigenic system has been well documented using alloantisera. We have produced the first mouse hybridomn cell line secreting an antibody. H1S41, which binds to leucocytes of rat strains carrying the RT7.2 but not the RT7.1 determinant. An lgG2b isotype switch variant (HIS4l.2b) of the original HIS41 (IgG1 isotype) was also made. HIS41 showed a clear and discrete binding in immunofluorescenl and histological experiments and has already been used in several studies on haemopoietic cell turnover and differentiation employing PVG rats congenic for RT7. The present study addresses the question of whether the RT7 gene products are members of the L‐CA family, which has been a matter of controversy over the last decade. When using HIS41 for the analysis of tissue distribution and molecular weight of RT7 gene products, a strong similarity was evident with the data reported for the L‐CA detected by MRC OX‐1 and MRC OX‐30. These two MoAb have been reported to bind to all members of the L‐CA family. All haemopoietic cells, excluding erythrocytes and the more mature stages of erythropoiesis, stained with HIS41. The molecular weights of HIS41 binding molecules on thymocytes and peripheral T cells were comparable to the L‐CA precipitated by MRC OX‐1. Capping and sequential immunoprecipitation studies indicated that HIS41 and MRC OX‐30‐ binding molecules were identical. MRC OX‐1. however, appeared to bind only a subset of these molecules. Thus, our study confirms the identity of RT7.2 gene products and L‐CA. lt also revealed a difference between MRC OX‐1 and MRC OX‐30 not noticed previously.
T‐Cell Activation. III. Attempts to Activate MHC Class I‐Negative and Class I‐Transfected EL4 T‐Lymphoma Cells by Immobilized Anti‐CD3 Antibody
Authors: CLAESSON M. H. , DISSING S. , PLESNER T. , STURMHÖFEL K. , BRAMS P.
Published: June 2006

The aim of this study was to examine whether the unresponsiveness of MHC class I‐negative subclones of the EL‐4 thymoma lo CD3 cross‐linking can be restored by transfection of class I genes into the H‐2‐negative cells. Cell activation experiments with selected MHC class I‐negative subclones and H‐2b‐and H‐2Ld‐positive transfectants showed that these cells are equally capable of secreting interleukin 2 (IL‐2) after exposure to the phorbol ester phorbol 12‐mvristalc 13‐acetale (PMA) and ionomycin. In contrast, only the parental H‐2‐positive EL4 cells are capable of responding to treatment with immobilized anti‐CD3 antibody with IL‐2 secretion and IL‐2 receptor expression. Measurements of intracellular free Ca2+ (Ca2+i) following anti‐CD3 antibody‐induced cross‐linking of parental EL4 cells and H‐2‐negativeand H‐2b gene‐transfected subclones showed that the parental cells and two of the class I transfectants, one H‐2‐positiveand one H‐2‐negative. responded with a slow rise in Ca2+,. whereas one H‐2‐positive transfected cell clone was completely refractory to CD3 cross‐linking. Modulation experiments using parental EL 4 cells, H‐2‐negative subclones and H‐2‐positive transfectants demonstrated that the CD3 and class I molecules of these different cells are modulated to the same extent after exposure lo specific antibodies. The present findings thus indicate that the unresponsiveness of H‐2‐negative EL4 subclone cells not CD3 cross‐linking is not functionally associated with a lack of class I surface expression.
Ciclosporin‐Induced Immunosuppression in Vitro
Authors: POVLSEN J. V. , RASMUSSEN A. , MADSEN M. , LAMM L. U.
Published: June 2006

The purpose of the present study was to analyse and correlate variations in lymphocyte sensitivity to, and binding of, ciclosporin (CsA) in vitro. Peripheral blood lymphocytes from healthy individuals were harvested over a 5‐week period and activated with purified protein derivative (PPD) or alloantigens in the presence or absence of CsA [I μg/ml). Sensitivity to CsA was expressed as the ability of the drug to suppress cell proliferation ([3H]thymidine incorporation) and high‐affinity imerleukin‐2 receptor (IL‐2R) expression. Binding capacity was tested in a [3H]CsA binding assay.

A significant variability in both sensitivity and binding capacity was recorded between individuals (P < 0.001) There was no correlation between high sensitivity and high binding capacity. The intraindividual day‐to‐day variability did not differ significantly from the experimental (intra‐ and interassay) variability. The CsA‐induced suppression of high‐affinity IL‐2R expression varied between 57.1 and 98.9%, while suppression of [3H]thymidine incorporation varied between 81.0 and 97.4% Specific binding of 10 nM[3H]CsA at 37° C varied between 5.4 and 10.7%.
Phenotype, Ultrastructure, and Function of CD1+DR+ Epidermal Cells that Express CD36 (OKM5) in Cutaneous T‐Cell Lymphoma
Authors: LISBY S. , BAADSGAARD O. , COOPER K. D. , HANSEN E. R. , MEHREGAN D. , THOMSEN K. , ALLEN E. , VEJLSGAARD G. LANGE
Published: June 2006

This study investigated the phenotype and function of different antigen‐presenting cells (APC) present within the epidermis of patients with cutaneous T‐cell lymphoma (CTCL). Involved epidermis of CTCL compared with uninvolved was found to contain increased numbers of CDI + DR+ APC. This population was heterogeneous and comprised both leucocytes of a novel CD1+DR+CD36 (OK.M5)+ phenotype and CD1+DR+CD36− indeterminate/Langerhans cells. The CD1+DR+CD36+ leucocytes did not express TcR‐1, CD5, CD 15. or CD22, and only a minor population expressed CD11, demonslrating that they were neither T nor B cells, and did not belong lo the major CD11+ (OKM1+) blood monocyte population. Electron microscopy of purified CD36+ lesional epidermal cells (EC) demonstrated that they lacked Birbeck granules found on CD1+‐selected Langerhans cells, and most cells exhibited features of indeterminate cells or macrophages.

The capacity of EC from involved epidermis to present alloanttgens was found to be increased relative to uninvolved epidermis in all patients tested, and this capacity was critically dependent upon the presence of CD45+ DR+ bone marrow‐derived cells but not on the presence of CD45+ DR+ keratinocytes. Positive selection using MoAb against CDI and CD36 demonstrated that both cell populations exhibited the capacity to stimulate T cells. The results indicate that a novel antigen‐presenting cell population with a unique phenotype is present within involved skin of patients with mycosis fungoides. These cells express CD36 in addition to CD1 and have an ultrastructural appearance consistent with a dendritic antigen‐presenting cell derivation.