bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunoelectrophoretic Characterization of Insulin Monomers and Polymers
Authors: KRØLL J.
Published: July 2006

Insulin monomers and polymers were analysed by quantitative immunoelectrophoretic procedures. The Zn‐insulin hexamer dissociated reversibly by dialysis against the Zn‐free electrophoresis buffer. The Zn‐insulin polymers showed precipitin reactions of partial identity. Monomeric salt‐free insulin migrated as soluble immune complexes in the antiserum gel. The insulin monomer did not absorb the precipitating antibodies against the Zn‐insulin polymers. Thus the polymer structure creates antigenic epitopes absent from the insulin monomer. As insulin is probably released from the β cells in the relatively stable form of Zn‐insulin hexamers, selective monomer assays might underestimate the total content of immunoreactive insulin in the biological fluids. Electroimmunoassay of Zn‐insulin immunoreactive antigens in human urine defines a normal reference range of 10–25 ng/ml.
Human B‐Cell Clones Expressing Lupus Nephritis‐Associated Anti‐DNA Idiotypes are Preferentially Expanded without Somatic Mutation
Authors: HTRABAYASHI Y. , MUNAKATA Y. , TAKAI O. , SHIBATA S. , SASAKI T. , SANO H.
Published: July 2006

Human monoclonal anti‐single/double‐stranded (ss/ds) DNA anlibodies (NE‐1 and NE‐13) expressed cross‐reactive idiotypes (Id), NE‐1 Id, which have been delected on the lupus glomeruli‐deposited anti‐DNA antibodies. The nucleotide sequences of the variable regions of NE‐1 and NE‐13 clones were analogous except for one nucleotide difference in the Vk region. The VH and Vk gene segments of NE‐13 clone were identical with germline genes VH4.21 and Vb(or Vb′). respectively. CDR3sof NE‐l and NE‐13 heavy chains were arginine rich and CDR1s contained an amino acid stretch. SGYY, the inverted sequence of YYGS. which was shared among CDR3s of several anti‐DNA antibodies. Clonal frequency analysis using a limiting dilution method revealed that NE‐1 Id‐positive clones at precursor cell level increased in lupus patients. These findings suggest that some IgM anti‐DNA clones which express NE‐1 Id associated with lupus nephritis use germline genes without mutation and they may be preferentially expanded at the precursor cell levels as well as at the mature cell level.
Evidence for an Ongoing Role of Class I Histocompatibility Molecules for the Production of Interleukin‐2 in Response to Suboptimal Concentrations of Phytohaemagglutinin
Authors: HASHIMOTO S. , McCOMBS C. C. , MICHALSKI J. P.
Published: July 2006

To examine the role of cellular interactions involving class I histocompatibility antigens in the response to low concentrations of phytohaemagglutinin, we studied the effect of antibodies to components of these antigens on proliferative responses, interleukin‐l and interleukin‐2 production, and IL‐2 receptor expression. Antibody to human β2‐microglobulin (β2m) had an inhibitory elTect both on IL‐2 accumulation at 48 h of culture and on the proliferative response 24 h later. Exogenous IL‐2 completely reconstituted the inhibited proliferative responses, and also restored the modest decrease in IL‐2 receptor expression that was induced by anli‐βm. Pretreatment of either purified monocytes or T cells with anti‐β2m had a simiiar inhibitory effect both on proliferation and on interleukin‐2 production. By contrast. IL‐1 production by LPS‐ or silica‐stimulated monocytes was not affected by this antibody. Kinetic experiments demonstrated that anti‐β2m was equally inhibitory when added at the initiation of culture or after 24 h. and significant inhibition occurred when the antibody was added as late as 48 h. Our results are consistent with an ongoing role for class I antigens in the cellular interactions between lymphocytes and accessory cells required for the production of IL‐2.
Cyclosporin A does not Inhibit IL‐lα‐Induced Epithelial Cell IL‐6 Secretion
Authors: HEDGES S. , AGACE W. , SVENSSON M. , SVANBORG C.
Published: July 2006

Trauma and infection activated a murine mucosal IL‐6 response in different ways: the IL‐6 response to bacteria was sensitive to Cyclosporin A (CsA); the IL‐6 response to trauma was not. The aim of the present study was to identify possible activators of the CsA‐insensitive IL‐6 secretion at the epithelial cell level. Two human epithelial cell lines from the kidney (A498) and bladder (J82) were exposed to Escherichia coli Hu734, interleukin‐lα (IL‐lα) and tumour necrosis factor a (TNF‐α). The E. coli strain had been used for the in vivo experiments which led to this study, and IL‐lα and TNF‐α were likely to be released during infections and trauma. The secretion of IL‐6 into the supernatants was compared between cells stimulated in the presence or absence of CsA. E. coli Hu734, IL‐lα and TNF‐α stimulated an IL‐6 response in the two epithelial cell lines. The IL‐lα‐induced IL‐6 response was rapid, and the secreted IL‐6 levels were significantly higher than those induced by E. coli Hu734 or TNF‐α. The IL‐6 response to IL‐ lα was insensitive to CsA. By contrast, the IL‐6 response to E. coli Hu734 and TNF‐α was inhibited by CsA. These results demonstrated that the inhibitory effect of CsA depends on the stimulus triggering the IL‐6 response. IL‐lα may play a role in the induction of trauma‐associated CsA‐insensitive IL‐6 secretion.
T‐Cell Immunity to Acetylcholine Receptor and its Subunits in Lewis Rats over the Course of Experimental Autoimmune Myasthenia Gravis
Authors: WANG Z.‐Y. , LINK H. , HUANG W.‐X.
Published: July 2006

Lymph nodes, spleen and thymus obtained from Lewis rats were examined over the course of experimental autoimmune myasthenia gravis (EAMG) for the distribution and the number of antigen‐reactive CD4+ T helper cells which, upon recognition of Torpedo acetylcholine receptor (AChR) or the α, β, γ or δ subunits of Torpedo AChR, responded by secretion of interferon‐gamma (IFN‐γ). T cells with these specificities were detected in these three immune organs. Numbers were highest in lymph nodes. In spleen and thymus, numbers of antigen‐reactive T cells did not differ. T cells reacting against the intact AChR were more frequent than T cells recognizing any of the subunits. The immunogenicity between the four subunits did not differ, with the exception that the α subunit induced a slightly higher T‐cell response. No restriction of the T‐cell repertoire to the four subunits was detected during early compared to late phases of EAMG. The AChR and subunit‐reactive T cells could—via secretion of effector molecules including IFN‐γ—play an important role in the initiation and perpetuation of EAMG. and consequently also of human myasthenia gravis. T cells with the same specificities were also detected in control animals injected with adjuvant only, but at much lower numbers which were within the range of T cells recognizing the control antigen myelin basic protein. They could represent naturally occurring autoimmune T cells.
Characterization of the Joining Chain (J‐Chain) Promoter
Authors: SIGVARDSSON M. , OLSSON L. , HöGBOM E. , LEANDERSON T.
Published: July 2006

A 300 b.p. promoter from the mouse joining chain (J‐chain) gene was studied with regard to functional activity and protein/DNA interactions. The promoter only stimulated expression of a chloramphenicol‐acetyl‐transferase (CAT) reporter gene when an enhancer was present in the construct, regardless of whether the construct was transfected into cell lines that did or did not express an endogenous J‐chain. Furthermore, deletion mutants lacking the 5’portion of the promoter were transcribed at a higher rate than the intact promoter in both J‐chain positive and J‐chain negative B‐cell lines but not in untransformed B lymphocytes stimulated by lipopolysaccharide, indicating the presence of a negative control element in the 5′ portion of the J‐chain promoter active in tumour cells only. The octamer element in the J‐chain promoter was found to bind Oct proteins, albeit with a low affinity. The penta‐deca (p.d.) element in the J‐chain promoter bound proteins in extracts from untransformed B cells but not in the tested cell lines. The protein binding to the J‐chain p.d, element did not compete efficiently with a. p.d. element from the SP6 k promoter. A protein binding to the 5′ portion of (he J‐chain was expressed in some cell lines but not in others; neither a negative nor a positive correlation to J‐chain expression could be seen. It was concluded that the J‐chain promoter is equivalent to a k promoter and that differentiation‐specific J‐chain expression is governed by distal, positive control elements located outside the analysed region.
Differential Expression of Rabbit Serum Amyloid A Genes in Response to Various Inflammatory Agents
Authors: RYGG M. , HUSBY G. , MARHAUG G.
Published: July 2006

Serum amyloid A (SAA) is an acute‐phase plasma protein which increases up to 1000‐fold after an acute‐phase stimulus. Several SAA genes and corresponding protein isotypes exist in individual species. Liver is the main source of production, but extra‐hepatic SAA expression has been described. In this study inflammation was induced in rabbits with lipopolysaccharide, turpentine, or casein. Transcription of SAA mRNA was studied using Northern blot analysis with probes specific for three different rabbit SAA isotypes and analysed by scanning densitometry. In the stimulated liver slight variation in SAA mRNA transcription level was seen after stimulation with different inflammatory agents. After lipopolysacchar‐ide‐stimulation SAA gene expression was also seen in most of the extra‐hepatic organs. After turpentine stimulation SAA mRNA was seen in the liver, the ovary, and the small intestines, and after casein stimulation only in the liver and the ovary. SAA1 and SAA2 were induced exclusively in the liver, while SAA3 was induced mainly in the extra‐hepatic organs. This indicates that the SAA genes probably are independently regulated both in relation to stimulus, gene‐ and tissue‐specificity.
Impaired Phagolysosomal Fusion of Peripheral Blood Monocytes from HIV‐Infected Subjects
Authors: PITTIS M. G. , STERNIK G. , SEN L. , DIEZ R. A. , PLANES N. , PIROLA D. , ESTEVEZ M. E.
Published: July 2006

We evaluated phagolysosomal fusion in peripheral blood monocytes from 20 HIV‐infected individuals and 40 normal controls, using a fluorescence assay with acridine orange as marker. The percentages of phagolysosomal fusion of monocytes from HIV‐infected subjects, after 30 and 60 min of yeast ingestion, (mean ± standard deviation) 57·2 ± 17 and 63·2 ± 18·6, respectively, when compared to normal controls (72·4 ± 7·8 and 77 ± 8·1), did not differ significantly. However, there was a direct linear association between the percentages of phagolysosomal fusion and CD4+ lymphocytes (P<0·001) or CD4/CD8 T‐cell ratio (P<0·01). These results suggest that phagolysosomal dysfunction becomes evident at late stages of HIV infection and progresses as CD4+ T‐lymphocyte count and CD4/CD8 T‐cell ratio decrease. On the other hand, recombinant gp120 inhibited significantly normal phagolysosomal fusion at concentrations ranging between 1 and 1000 ng/ml. Taking together the results obtained, we can conclude that gp120 could be responsible for monocyte phagolysosomal dysfunction observed in HIV infected patients.
Endothelial Cells Promote Anti‐CD3‐Induced T‐cell Proliferation via Cell‐Cell Contact Mediated by LFA‐1 and CD2
Authors: WESTPHAL J. R. , WILLEMS H. W. , TAX W. J. M. , KOENE R. A. P. , RUITER D. J. , DE WAAL R. M. W.
Published: July 2006

T‐cell activation requires not only T‐cell receptor (TCR) engagement and subsequent TCR/CD3 cross‐linking, but also one or more secondary activation signals generated by accessory cells (AC). We investigated the accessory function endothelial cells (EC) in an in vitro model for T‐cell activation where the first cross‐linking signal was delivered to peripheral human T lymphocytes by cither immobilized anti‐CD3 monoclonal antibody (MoAb) or by PHA. In a previous report, we showed that EC provided a potent costimulatory signal in this model system. We have now analysed the nature of the EC‐derived costirnulatory signal by testing whether EC could be substituted by cy lokines. by studying the effect of EC fixation and by testing the involvement of a number of adhesion molecules. Our findings indicate that EC accessory function is mediated mainly by membrane‐bound factors. The nature of these membrane‐bound factors was analysed by studying the inhibitory properties of a series of MoAbs directed against several adhesion molecules. Antibodies directed against CD44, E‐selectin, CD31, CD26, B7/BBI, VLA‐4 or VCAM‐1 were not inhibitory. However, an inhibition, was clearly observed with antibodies against LFA‐1 and CD2. Remarkably, this inhibition was not found with MoAbs to their respective counterstructures ICAM‐1 and LFA‐3. In summary, we postulate that both LFA‐l/ICAM‐1, and CD2 LFA‐3 interactions are involved in EC accessory function, although the role of the EC‐associated adhesion partners is not fully understood.
Serum Amyloid‐P Component of the Armenian Hamster: Gene Structure and Comparison with Structure and Expression of the SAP Gene from Syrian Hamster
Authors: RUDNICK C. M , DOWTON S. B.
Published: July 2006

Serum amyloid P(SAP), a phylogenetically conserved pentraxin, is an integral component of all amyloid deposits. Regulation of expression of SAP gene expression is quite different in two related hamster species. In Syrian hamsters, the resting serum levels of SAP are determined by gender, and the direction of alteration following inflammation is divergent. In Armenian hamsters, SAP is not a prominent acute‐phase reactant and there is no gender dimorphism of expression.

The structure and expression of the SAP gene of the Armenian hamsters was investigated by isolation of genomic clones, nucleotide sequence analysis, and RNA studies. The gene structure of Armenian hamster SAP is similar to the genes of all other pentraxins studied. While the upstream regions of the SAP genes of Syrian and Armenian hamsters are quite similar, important differences in potential enhancer sites have been recognized by comparing the corresponding sequences of SAP genes from both species. Little alteration in hepatic levels of transcripts encoding SAP or CRP, the other pentraxin, were noted following administration of lipopolysaccharide to Armenian hamsters. This relative lack of response occurred despite a marked acute phase reaction documented for serum amyloid A mRNA levels.
Analysis of the Neutralizing Antibody Response to the Measles Virus Using Synthetic Peptides of the Haemagglutinin Protein
Authors: MULLER C. P. , SCHROEDER T. , TU R. , BRONS N. H C. , JUNG G. , SCHNEIDER F. , WIESMÜLLER K. H.
Published: July 2006

Infection or immunization with measles virus induces a protective immune reaction including neutralizing antibodies against the haemagglutinin and fusion protein. The reactivity of the polyclonal IgG response of sera obtained from late convalescent donors was studied, using overlapping 15mer peptides covering the complete sequence of the measles virus haemagglutinin. Most sera reacted with a similar set of peptides generating a characteristic binding pattern. The reactive peptides correspond to a region mediating cell hemolysis (aa310–325), to regions which serve as targets to neutralizing antibodies and to a putative transmembrane region (aa35–58). The latter region contains also a human T‐cell epitope providing evidence of a non‐random association of T‐ and B‐cell epitopes. We also immunized different strains of mice and rabbits with measles virus. In contrast to the human sera, animal sera with strong neutralizing activities did not react with any of the H‐protein peptides. The mostly weak reactivities with the linear sequences contrast with the strong neutralizing activities of the human or animal antibodies, suggesting that these primarily recognize the fusion protein or conformational epitopes of the haemagglutinin protein.
Bacterial Lysates and Ribosomes as Inducers of Specific Immune Responses: a Comparative Study
Authors: BéNé M. C , KAHL L. , PERRUCHET A. M. , HERMES H. , MOSGES M. , NORMIER G. , BINZ H. , FAURE G. C.
Published: July 2006

A bacterial lysate (OM‐85 BV), a preparation of purified bacterial ribosomes (D53) and a placebo were tested for ability to induce the local appearance of specific antibody‐containing cells. The three compounds were given orally to 90 children who required tonsillectomy. Surgery was carried out after 1 month of therapy. Frozen‐cut sections of each tonsil were tested in indirect immunofluorescence. Cells containing antibodies directed to Streptococcus pneumoniae. Streptococcus pyogenes. Haemophilus influence or Klebsiella pneumoniae were enumerated. Lowest values were noted in the placebo group. Slightly higher numbers were observed after treatment with OM‐85 BV, but significant increases were noted only for the elevated numbers of specific antibody‐containing cells observed after D53 therapy. Bacterial ribosomal preparations thus contribute efficient induction of specific local immune responses in man.
Human Naive and Memory T‐Helper Cells Display Distinct Adhesion Properties to ICAM‐1, LFA‐3 and B7 Molecules
Authors: PARRA E. , WINGREN A. G. , HEDLUND G. , SJÖGREN KALLAND H.‐O. , SANSOM D. , DOHLSTEN M.
Published: July 2006

In this paper the contribution of different accessory molecules to the adhesion of resting, naive and memory CD4+ T cells was examined utilizing a panel of CHO cell transfectants as model antigen‐presenting cells (APCs). CD4+ T lymphocytes demonstrated strong adhesion to HLA‐DR4 transfected CHO cells co‐expressing B7, ICAM‐I or LFA‐3 molecules, suggesting that all three adhesion pathways is utilized by resting CD4+ cells. Monoclonal antibodies (MoAbs) against the corresponding receptors on T cells, e.g. anti‐CD28, anti‐LFA‐1β and anti‐CD2, inhibited completely T‐cell adhesion to natural ligands expressed on transfected CHOcells. Pretreatment of CD4+ T cells with NKI‐L16 MoAb, which interact with an activation epitope on LFA‐loc chain, enhanced adhesion to ICAM‐1 but not B7 or LFA‐3 expressing CHO cells. Analysis of T helper‐cell subsets revealed that memory T cells bound several fold stronger to ICAM‐1 expressing transfectants compared to the CD4+ 45RA+ naive T cells, whereas adhesion to B7, LFA‐3‐ and B7/LFA‐3‐expressing CHO cells was similar in both T‐cell subsets. The kinetics of adhesion of naive and memory CD4+ T cells to ICAM‐1 was rapid and similar in both subsets. The NKI‐L16 MoAb multiplied several times ICAM‐1‐dependent adhesion in naive compared to memory cells, which enabled the naive cells to reach a similar adhesion level as memory cells. The results suggest that resting naive CD4+ T cells utilize preferentially the CD2/LFA‐3 or CD28/B7 adhesion pathways upon adhesion to APCs, while memory CD4+ T cells utilize the CD2/LFA‐3, CD28/B7 and LFA‐l/ICAM‐1 adhesion pathways. The NK.I‐L16 MoAb‐induced upregulation of adhesion involves an increased affinity of LFA‐1 for its ligand and not a change in the number of LFA‐1 molecules. This is compatible with a view that naive cells express a large number of inactive LFA‐1 molecules, whereas memory cells express preferentially activated LFA‐1 molecules. The inherent low number of active LFA‐I molecules on naive CD4+ T cells may be important in keeping these cells in a resting state.
Redistribution of Lymphocytes Following E. coli Sepsis
Authors: TOFT P. , LILLEVANG S. T. , TØNNESEN E. , SVENDSEN P. , HöHNDORF K.
Published: July 2006

Infusion ofendotoxin elicits lymphopcnia and a transient granulocytopcnia followed by granulocytosis in peripheral blood. The purpose of this study was to investigate which tissues the lymphocytes are redistributed to in response to endotoxaemia. Lymphocytes were isolated from the peripheral blood of 20 rabbits, labelled with III lndium‐tropolene and reinjected intravenously into the rabbits. Ten rabbits received an infusion of Escherichia coli endotoxin 2μg/kg‐1, while 10 rabbits received isotonic saline and served as a control group. The redistribution of lymphocytes was imaged with a gamma camera, and calculated with an interfaced computer before, and 2, 4 and 6 h after infusion of endotoxin or saline. Interleukin‐lB and serum cortisol were measured. Following endotoxaemia the lymphocytes in peripheral blood decreased from 1.95 109/1 to 0.83 6 h later. Interleukin‐lB and serum cortisol increased significantly. The radioactivity of labelled cells in the spleen and in the heart and lungs decreased to 83.3% and 87.8% of initial values respectively, 6h after infusion of endotoxin. The radioactivity of the lymphatic tissue in and around the intestine increased to 128.8% of initial values. The results indicate that endotoxaemia induces redistribution of lymphocytes from peripheral blood and spleen to lymphatic tissue.
Quantitative Analysis of Lymphocyte CD1 la using Standardized Flow Cytometry
Authors: PALLIS M. , ROBINS A. , POWELL R.
Published: July 2006

CDlla is the alpha‐subunit of the leucocyte adhesion and costimulation molecule LFA‐1. We have refined the measurement of lymphocyte CDlla density with a FACScan using commercially available fluorescent beads for standardization and fluorescein‐conjugated antibody to CDlla of known fluorescein:protein ratio. The fluorescence intensity of CDlla on peripheral blood CD4+ and CD8+ lymphocytes was measured in 60 healthy subjects. We demonstrated linear correlation between age and mean CDlla density (r =0.47 for CD4+ cells, r = 0.71 for CD8+ cells). We established that there is a consistent logical cut off point at 4.3 × 103 bound antibody molecules between low‐expressing and high‐expressing subsets of CD8+ cells and we then investigated whether the variation in lymphocyte CDlla expression in healthy subjects was sufficiently small for the application of this method to the detection of abnormal groups or individuals. Analysis of the CDlla high subsets has high statistical power (>99% in 60 subjects to detect a 25% difference) and good precision (<4% differences). The advantages of the method for comparative studies of cell surface accessory molecules are discussed. We have also evaluated a frozen cell line for quality control, and demonstrated up‐regulation of CD1la density on CD4+ and CD8+ cells measured in three patients with infectious mononucleosis.
B Cells Carrying Surrogate Receptors in their Membranes Process and Present Antigen to Specific Murine T Cells
Authors: GONTIJO C. M. , MÖLLER G.
Published: July 2006

A palmitate‐conjugate derivative of ovalbumin which can be inserted into the membrane of B cells has been prepared. The ability of these cells to act as antigen‐presenting cells for specific T lymphocytes obtained from immunized mice was tested. It was found that the conjugates were more efficiently processed and presented than the naive form of the antigen.

Palmitate‐conjugated antibodies specific to ovalbumin were also inserted into the cell membrane of normal B lymphocytes. These cells were pulsed with the antigen and tested as antigen‐presenting cells for T cells obtained from immunized mice. The antibody‐decorated B cells presented ovalbumin more efficiently than non‐decorated controls.

Whether antibody‐decorated, antigen‐pulsed B cells could prime T cells in viro was investigated. Some priming activity was found.
Peptide Binding Sites Recognized by Anti‐Mucin (MUC2) Monoclonal Antibodies
Authors: XING P.‐X. , APOSTOLOPOULOS V. , PRENZOSKA J. , DEVINE P. L. , McKENZIE I. F. C.
Published: July 2006

Multiple genes coding for human mucins have been identified (MUC1‐5) and here monoclonal antibodies (MoAb) to a gastrointestinal mucin‐MUC2 are examined. The antibodies were made to a synthetic peptide representing a single repeat in the core protein of the variable number of tandem repeat region. Using the six‐mer overlapping peptides synthesized on polyethylene pins, different binding sites were detected by five anti‐MUC 2 MoAbs. These contained amino acids: STTT, PTT, GTQTP, TPTP and PTTT (one antibody), and TPTPT. The repeat region of MUC2 essentially is hydrophobic, but contain useful immunogenic sites. This information will be useful for studying the structure and function of MUC2.
Partial Biochemical Characterization and Purification of IgG2b Inducing Factor as a New Cytokine from Synovial Fluid of Patients with Rheumatoid Arthritis
Authors: ABEDI‐VALUGERDI M. , RIDDERSTAD A. , STRÖM H. , MÖLLER G. , MOLLER E.
Published: July 2006

Rheumatoid arthritis synovial fluid (RA‐SF) contains a novel biological activity, which selectively induces IgG2b antibody production in lipopolysaccharide (LPS)‐activated mouse spleen cells in vitro and in vivo. Our previous studies have shown that this activity is not functionally identical to otherwell‐known cytokines and interleukins. In this study we demonstrate the partial purification and biochemical characterization of the IgG2b inducing activity in RA‐SF. Biochemical characterization revealed that the IgG2b inducing activity in RA‐SF has the following properties: it is a protein, sensitive to pH > 11 and < 4, which is precipitated by 50% of saturated ammonium sulphate and has a molecular weight of 50–70 kDa; it binds to Cibacron‐blue and heparin and its activity is not mediated by immunoglobulins or immune complexes, which are present in RA‐SF. Biochemical characteristics of the IgG2b inducing activity also differ from other cytokines and interleukins. The term IgG2b inducing factor is proposed for this novel activity.
Epitope‐Specific Differential Modulations within the T‐Cell Antigen Receptor Complex
Authors: SUZUKI S. , TOKUDOMÉ T. , YACHELINI P. , SAIZAWA M. K.
Published: July 2006

The feeding of cloned helper T cells by irradiated spleen cells results in the modulation of T‐cell antigen receptors (TCRs) as detected by FACS. When two groups of anti‐TCR antibodies—anti‐ldiotypic and anti‐Vβ—are compared, time‐dependent patterns of modulation of TCR are drastically different. Whereas up to 80% of the idiotypic determinants disappear within 2 days after stimulation, almost no change is observed for the determinants recognized by Vβ‐specific antibodies within the same period. Sequential immunoprecipitations clearly showed that anti‐idiotypicand anti‐Vβ antibodies recognize the same population of TCR. Moreover, surface labelling followed by immunoprecipitation of TCR did not show rapid and drastic changes in the amount of specifically precipitated material. The modulation is not significantly affected by the addition of specific antigen, but is specific for the clone's MHC restriction element. Functionally, the modulation of TCR epitopes affects the ability of anti‐TCR antibody but not of the MHC/antigen complex to stimulate T cells. We conclude that the observed modulation of fluorescence intensity in the TCR complex after MHC recognition (1) is epitope specific. (2) is independent of the presence of nominal antigen and (3) does not affect the efficiency of antigen‐specific stimulation.
The Primary Structure of Rabbit Serum Amyloid A Protein Isolated from Acute Phase Serum
Authors: SYVERSEN P. V. , JUUL J. , RYGG M. , SLETTEN K. , HUSBY G. , MARHAUG G.
Published: July 2006

Serum amyloid A (SAA) protein, a sensitive acute phase protein and the precursor of protein AA in secondary amyloid, was purified from pooled acute phase rabbit serum using two different methods: isolation of protein SAA directly by octyl‐Sepharose chromatography of total serum, and dissociation and isolation of apoSAA from acute phase high density lipoprotein (HDL). The protein SAA fraction obtained was further purified using gel filtration and ion exchange chromatography.

Rabbit protein SAA has 104 amino acid residues, like human SAA, and has a partially blocked N terminus. The highly conserved region from position 33 to position 63 found in SAA from all species studied was confirmed also in rabbit SAA. No microheterogeneities were observed.

The amino acid sequence showed extensive N‐terminal homology with the rabbit amyloid A protein, except for the microheterogeneity in position 12 in protein AA. It also showed identical amino acid sequence with that deduced from the rabbit cDNA clone pSAA 55. Complete homologies were found with clone SAA 2, except for positions 22 and 78, clone SA8‐1, except for positions 22 and 79 and clone SA7‐3, except for position 22. This pSAA 55/SA7‐3/SA8‐l/SAA2‐like protein was the only SAA isotype found both in total serum and in the HDL fraction. Isotypes corresponding to other SAA‐like genes could not be found in this pool of acute phase rabbit sera.
Cholera Toxin Enhances Alloantigen Presentation by Cultured Intestinal Epithelial Cells
Authors: BROMANDER A. K. , KJERRULF M. , HOLMGREN J. , LYCKE N.
Published: July 2006

In the present study we show that cholera toxin (CT) strongly potentiates antigen presentation by intestinal epithelial cells, probably by enhancing co‐stimulation. This was demonstrated in an allogeneic system using cells from the IEC‐17 rat epithelial cell line as antigen presenting cells (APC). These cells were induced by optimal concentrations of IFN‐γ to express good amounts of Ia antigen and cultured for 24–48 h in the presence or absence of CT. Thereafter the cells were thoroughly washed and added to cultures containing MHC‐incompatible spleen cells as responder cells. Epithelial cells exposed to CT demonstrated greatly enhanced ability to trigger allogen‐specific T‐cell proliferation as compared with IEC‐17 cells treated with IFN‐γ alone. The mechanism for the enhanced APC function was investigated by analysing CT‐treated IEC‐17 cells for increased class II MHC antigen expression or enhanced production of cytokines with known co‐stimulatory function. We found no significant increase in class II MHC antigen expression. By contrast, CT strongly promoted, in a dose‐dependent fashion, the production of both IL‐1 and IL‐6 cytokines by IEC‐17 cells as compared with untreated epithelial cells. This effect of CT was specific and not due to contaminating endotoxin because excess amounts of soluble toxin receptor, ganglioside GM1, added to the IEC‐17 cultures completely abrogated the cytokine response to CT.

These results together with our previous findings of enhanced antigen presentation by macrophages stimulated by CT suggest that the potent adjuvant function of CT for induction of mucosal immune responses might be attributed to an enhanced co‐stimulating ability of several putative APC in the mucosal immune system: macrophages, B cells and epithelial cells.
A Low Serum Concentration of Mannan‐Binding Protein is Not Associated with Serogroup B or C Meningococcal Disease
Authors: GARRED P. , MICHAELSEN T. E. , BJUNE G. , THIEL S. , SVEJGAARD A.
Published: July 2006

The mammalian C‐type serum lectin, mannan‐binding protein (MBP), may induce Clq‐ and antibody‐independent activation of the classical pathway of complement. Accordingly, MBP is considered as a member of the complement system. Complement deficiencies have been found with increased frequency in patients with meningococcal disease. Therefore, we investigated the MBP levels in patients with meningococcal disease. Ninety‐nine Norwegian individuals (age 12–21 years) who survived severe systemic disease caused by serogroup B or C meningococci were investigated. No significant differences were observed in the MBP concentration between patients with serogroup B (n = 76) or C (n = 25) disease and healthy blood donor controls (n = 40) (P >0.05). The frequency of patients with low levels of MBP (< 100 μg/1) was 10.1%. This was not different from controls (12.5%). Thus, low MBP concentrations do not appear to predispose to serogroup B or C meningococcal disease.
The IE Allogeneic Response of T Cells from C57B1/6 Mice is Associated with Genes in the TCRa Locus
Authors: RUBIN B. , GOUAILLARD C. , WIEDERANDERS G. , KUHLMANN J.
Published: July 2006

It has been demonstrated that induction of immune responses, infectious diseases and autoimmune manifestations can be associated with at least four gene loci: the major histocompatibility complex (MHC) locus; the immunoglobulin (Ig) heavy chain (Hc) locus; and the T–cell receptor (TCR) TCR–β or TCR–β chain loci. In the present study, we have analysed whether T–cell responses of IE–negative C57B1/6 (B6) mice to IE alloantigen (IEα transgenic B6 mice = B6. Eα 16) or to trinitrophenylated (TNP) syngeneic spleen cells were influenced by changes in the Ig–Hc locus or the TCRa locus. Whereas the fine specificity of T–cell responses to IE alloantigen was the same in B6 mice and in Ig–Hc congenic B6.26a or TCRa congenic B6.10TCa mice, the latter strain of mice demonstrated much higher IE–specific T–cell responses against B6. Eα16 spleen cells than B6 or B6.26a mice. This high responsiveness was a dominant feature and associated with the TCRa locus. In addition, the TCRVα or Vα repertoire of the congenic strains of mice was polyclonal and very similar. The TNP–specific T–cell responses of B6 and B6. 10TCa mice showed the same restricted TCRVα and Vα repertoire. It is concluded that in both an oligoclonal T–cell response (anti–TNP) and a polyclonal T–cell response (anti–IE), exchange of Ig–Hc or TCRa loci does not significantly influence the TCRVα or Vα repertoire in IE–negative C57B1/6 mice.
Low Prevalence of Complement Deficiencies among Patients with Meningococcal Disease in Norway
Authors: HØGÅSEN K. , MICHAELSEN T. , MELLBYE O. J. , BJUNE G.
Published: July 2006

Sera from 98 individuals who had survived meningococcal disease were analysed for classical and alternative pathway haemolytic activity and the complement components C3. C4 and properdin. No complete deficiency was found. However, median properdin concentration was only 86% in the disease group compared with the controls (P<0.001). Properdin was also significantly lower in serogroup C disease (median 76%.) compared with serogroup B disease (median 90%, P = 0.005). Severe properdin deficiency is an established risk factor for meningococcal disease. The present data may indicate that even moderately reduced properdin level can increase the risk of developing meningococcal disease.
Preparation and Reactivities of Anti‐Porcine CD44 Monoclonal Antibodies
Authors: YANG H. , HUTCHINGS A. , BINNS R. M.
Published: July 2006

Five monoclonal antibodies (MoAbs) were raised against porcine soluble CD44. The MoAbs recognized the same antigen on the surface of porcine lymphocytes as was recognized by anti‐human CD44 MoAb Hermes‐1, but identified five different epitopes. They bound to most porcine leucocytes but not to red cells. The epitopes were susceptible to treatment with papain or bromelain, whereas trypsinization of porcine leucocytes only reduced the antigen density. The epitopes seem to be co‐expressed among various lymphoid tissues.

The MoAbs also cross‐reacted to various degrees with leucocytes of humans, dogs, sheep, cattle, goats and horses, suggesting that the corresponding epitopes arc differentially conseted atnong species.
Linkage Disequilibrium within the HLA Complex does not extend into HLA‐DP
Authors: BAISCH J. M. , CAPRA J. D.
Published: July 2006

It is well known that certain alleles from different loci within the Human Leucocyte Antigen (HLA) complex are in linkage disequilibrium. This linkage phenomenon is relatively well characterized for haplotypes that include specific class I and class II alleles such as HLA‐B8 and HLA‐DR3. However, the HLA‐DP genes are located at the centromeric end of the HLA complex and are less well characterized with regard to linkage disequilibrium. The availability of a large population of healthy subjects and sequence‐specific oligonucleotide (SSO) typing enabled us to assess the degree of linkage between HLA‐DPBl and HLA‐DQBl genes. Using the polymerase chain reaction and a series of oligonucleotide probes which define seven DQβ alleles and twenty DPβ alleles, we studied 180 unrelated, normal Caucasian individuals and found only weak or negative associations between HLA‐DPBl and HLA‐DQBl. These data demonstrate that the association between HLA‐DQ and DP is weak and also imply that DP extended haplotypes related to particular diseases may not reflect normal associations. Implications of these results might impact on the concept of linkage disequilibrium in general as well as the evolution of the HLA complex. In addition, extensions of this work may have clinical ramifications with regard to bone marrow transplantation and founder effects in certain diseases.
Interferon‐Gamma Stimulates Neopterin Release from Cultured Kidney Epithelial Cells
Authors: MOUTABARRIK A. , TAKAHARA S. , NAKANISHI I. , KOKADO Y. , TAKANO Y. , KAMEOKA H. , ISHIBASHI M. , ZAID D.
Published: July 2006

The ability of cultured kidney epithelial cells (KEC) to secrete neopterin, which is a marker of the activation of immune system was studied. In this study inlerferon gamma (IFN‐γ) was shown to induce neopterin release from KEC in a dose‐ and time‐dependent manner. Many other cytokines and mitogens (IL‐1β, IL‐2, IL‐6, LPS and phorbol ester) were tested for their ability to induce neopterin in KEC but all failed to induce a significant release of neopterin from KEC. By itself TNF‐ á induced a release of small amounts of neopterin but strongly potentiated the effect of IFN‐γ in a synergistic manner to induce neopterin secretion. These data indicate that not only monocytes and macrophages which it is well known secrete neopterin, but KEC are responsible also for the high serum or urine level of neopterin observed in patients with kidney allograft rejection or infections episodes. As the amount of neopterin released by KEC is smaller than that secreted by activated macrophages, the contribution of KEC to the overall production of neopterin during certain diseases may be small.
Fc Receptors Expression and Function in Mononuclear Phagocytes from AIDS Patients: Modulation by IFN‐Gamma
Authors: CAPSONI F. , MINONZIO F. , ONGARI A. M. , BONARA P. , PINTO G. , CARBONELLI V. , LAZZARIN A. , ZANUSSI C.
Published: July 2006

Fc‐receptor (FcR)‐mediated phagocytosis and FcR (FcRI, FcRII and FcRIII) membrane expression was studied on freshly separated and cultured monocyles (Mo) from 20 AIDS patients and 20 healthy controls. Both Mo and Mo‐derived macrophages from AIDS patients presented a significant defect in their capacity to ingest IgG‐coated erythrocytes (EA) compared to control cells. This functional defect did not depend on a decline in the n umber of FcR+ cells or on a decrease in the expression of FcR on their surface. In fact, the percentages of phagocytes reacting with anti‐FcRI Mo Ab (32.2) or ami‐FcR II MoAb (IV. 3) were similar for controls and AIDS patients, while the percentage of FcRIII‐positive Mo (MoAb 3G8) was higher in the AIDS population than in controls, though this difference was not seen on cultured Mo. The level of FcRI expression, evaluated as mean fluorescence intensity (MFI), was higher on freshly separated Mo from AIDS patients than from controls but this difference disappeared also with differentiation of Mo to Mo‐derived macrophages in vitro. Parallel analysis of FcRII and FcRIII onphagocytes revealed no differences in the MFI between the AIDS and control groups. Some observations suggested that this functional defect might be secondary to phagocyte priming by circulating lFN‐γ: (1) in vitro stimulation of Mo with hrIFN‐γ, which increased FcRI expression, actually reduced phagocytosis of IgG‐coated particles; and (2) IFN‐γ concentrations were increased in AIDS patients' plasma. In spite of these findings, no significant correlation was found between plasma IFN‐γ concentrations and FcR‐mediated ingestion in AIDS patients, making the hypothesis uncertain. Even if the basis for the impaired FcR‐mediated phagocytosis in AIDS patients remains unclear, this functional defect may have a role in the immunopathogenesis of AIDS, constituting a component cause of the immunodeficiency.
The Interaction between Beta 2‐Microglobulin (ßm) and Purified Class‐I Major Histocompatibility (MHC) Antigen
Authors: PEDERSEN L. Ø. , HANSEN A. S. , OLSEN A. C. , GERWIEN J. , NISSEN M. H. , BUUS S.
Published: July 2006

The function of MHC class‐I molecules is to sample peptides from the intracellular environment and present them to CD8+ cytotoxic T lymphocytes. To understand the molecular details of the assembly (and disassembly) of peptide‐ß2m‐class‐I complexes a biochemical peptidc‐class‐I binding assay has been generated recently and this paper reports on a similar assay for the interaction between ß2m and class I. As a model system human ß2m binding to mouse class I was used. The assay is strictly biochemical using purified reagents which interact in solution and complex formation is determined by size separation. It is specific and highly sensitive. The observed affinity of the interaction, KD, is close to 0.4 nw. The rate of association at 37 C is very fasi (the ka is around 5 × 104/M/s) whereas the dissociation is slow (the kd is around 8 × 10−6/s); the ratio of dissociation to association yields a calculated KD close to the observed value. At 37° C almost all of the purified class I participates in binding of the exogenously offered ß2m showing that a considerable exchange of the endogenous ß2m occurs. Finally, it was demonstrated that exogenous ß2m enhances binding to MHC class‐I of short perfectly‐matching peplides as well as longer peptides.
Global Analysis of Antibody Repertoires. 1. An Immunoblot Method for the Quantitative Screening of a Large Number of Reactivities
Authors: HAURY M. , GRANDIEN A. , SUNDBLAD A. , COUTINHO A. , NOBREGA A.
Published: July 2006

This paper describes a procedure for analysing multiple antibody reactivities that explores a commercially available immunoblot system, and is based on a double staining of nitrocellulose membranes, revealing both antibody reactivities and the migration position of the blotted proteins in the membrane. Quantification of both stainings by densitometry allowed the accurate superposition of the immunoreactivity and total protein profiles of each Line. Moreover, the protein stainings ofthe different lanes could be adjusted with a simple‐scale transformation algorithm, correcting for possible distortions during electrophoretic migration, and allowing for the precise comparison ofthe immunoreactivity profiles in different lanes. The procedure is discriminatory enough to identify unique reactivity patterns in random pools of 104 activated B cells, and to define strain‐specific natural antibody repertoires. The utilily of this immunoblot method as an assay for simultaneously scoring multiple reactivities to hundreds of antigens in complex mixtures of antibodies, and thus defining antibody repertoires in a global manner, is discussed.
Utilization of a Potentially Universal Downstream Primer in the Rapid Identification and Characterization of Vβ Genes from Two New Human Vλ Gene Families
Authors: DEFTOS M. , SOTO‐GIL R. , QUAN M. , OLEE T. , CHEN P. P.
Published: July 2006

Polymerase chain reaction (PCR) has increased dramatically the speed of cloning and characterizing numerous genes. However, its application to identifying and analysing new germline Ig‐variable (V) gene families has been hampered by the lack of sequence information in the downstream flanking regions of the concerned V genes, which are deleted during V(D)J rearrangements. To circumvent this problem, the possibility was explored that a degenerate downstream primer may be used in conjunction with a specific upstream primer, to clone members of new Vβ gene families, as much less is known about Vλ genes than Vh and Vk genes in humans. Firstly the feasibility and the specificity of a degenerate primer was examined by comparing it with an established downstream primer in amplifying known Vλ1 genes. The results were positive. Thus, the degenerate primer was used to clone and characterize germline Vλ genes of the recently defined Vλ8 and Vλ9 gene families. This current strategy may help speed up the identification and characterization of all human Vλ genes. Moreover, a similar strategy can be applied to identify and characterize rapidly new V genes of either known or unknown Ig and T‐cell receptor V gene families.
Major Histocompatibility Complex Class‐II Alleles in Primary Biliary Cirrhosis
Authors: ZHANG L. , WEETMAN A. P. , BASSENDINE M. , OLIVEIRA D. B. G.
Published: July 2006

The major histocompatibility complex (MHC) class‐II alleles at the DRBI, DQBl and DPBl loci were investigated in 40 patients with primary biliary cirrhosis (PBC) and 43 local healthy controls. Restriction fragment length polymorphism (RFLP) was used for DRBI typing. DQBl and DPBl regions were amplified using the polymerase chain reaction (PCR) and then probed with 32P‐labelled allele‐specific oligonucleotide probes. There was an increased frequency of DR8 (10% compared to 4% in controls), and a decreased frequency of DR2 (18% compared with 28% in controls) in patients with PBC, but the differences were not significant. There were no significant differences for the other DR alleles, the seven DQ alleles, or the eight DP alleles. In conclusion, no significant MHC class‐II associations with primary biliary cirrhosis have been demonstrated.
Improvement of TH1 Functions during the Regulation Phase of Mercury Disease in Brown Norway Rats
Authors: CASTEDO M. , PELLETIER L. , PASQUIER R. , DRUET P.
Published: July 2006

Brown Norway (BN) rats are poor responders to T‐cell mitogens and alloantigens when compared to Lewis (LEW) rats. This is dependent partly upon a defect in IL‐2 production. The TH2‐mediated immune abnormalities observed in BN rats injected with mercuric chloride (HgCl2) are self‐limited and it is probable that this regulation phase involves TH1 ‐like cells. This paper reports on a study of the ability of lymph node cells (LNC) from normal BN and LEW rats and from HgCl2‐injected BN rats to produce IL‐2 and to proliferate when stimulated in vitro by Con A or alloantigens in mixed lymphocyte reaction (MLR), as well as to develop a cytotoxic T lymphocyte (CTL) response to alloantigens. This study will confirm that LNC from BN rats proliferate less than LNC from LEW rats, that the former produce less IL‐2 than the latter, and that the proliferative response is restored partially after addition of IL‐2. In addition, it is shown (1) that the CTL response is defective in normal BN rats when compared to that of normal LEW rats, and (2) that, after the second week of HgCl2 injections, the proliferative responses to Con A and alloantigens are improved as well as IL‐2 production, and a complete restoration of CTL function is observed. These results show that normal BN rats are deficient in the induction of THl‐like cells and that, from the second week of HgCl2 injections, these TH1 functions improve.
Influence of Tumour Necrosis Factor‐α on the Expression of Fc IgG and IgA Receptors, and Other Markers by Cultured Human Blood Monocytes and U937 Cells
Authors: GESSL A. , WILLHEIM M. , SPITTLER A. , AGIS H. , KRUGLUGER W. , BOLTZ‐NITULESCU G.
Published: July 2006

The expression of Fc receptors for IgG (FcγR) and IgA (FcαR) and of various other antigens on the human monocytic cell line U937 and peripheral blood monocytes, under stimulation with human recombinant tumour necrosis factor‐α (TNF‐α) and other cytokines, was investigated by flow cytomelry. TNF‐α, as well as interferon‐γ (IFN‐γ) or interleukin‐6 (IL‐6) had a significant up‐regulating effect on U937 expression of FcγRI/CD64. Furthermore, the action of TNF‐α was augmented by IL‐6, and more evidently by IFN‐γ. IFN‐α alone had only a marginal effect, but was able to increase the TNF‐α‐driven FcγRI expression. In contrast to U937 cells, TNF‐α did not enhance significantly FcγRI expression on human monocytes. Interestingly, on both U937 cells and monocytes. FcαR was augmented markedly by TNF‐α. Furthermore, TNF‐α induced the expression of HLA‐DR and HLA‐DP antigens on monocytes and U937 cells. The expression of FcγRII/CD32, FcγRIII/CD16. CD14, complement receptor type 1 (CR1/CD35). CR4 (CD11c/CD18), and MHC class‐I antigens, was not influenced significantly by TNF‐α. The results of this study show that TNF‐α may act on human mononuclear phagocytes, alone or in combination with other cytokines, by modulating the expression of various cell‐surface antigens.