bluuebunny/crossref_metadata_2025 huggingface dataset:
Bipolar heterothallism in B‐group isolates of Leptosphaeria maculans
Authors: SOMDA I. , HARKOUS S. , BRUN H.
Published: November 2003

Pairwise combinations were carried out between the eight isolates originating from each of three asci of the B‐group of Leptosphaeria maculans. Some of the pairings produced mature pseudothecia when incubated on wheat straws at 18°C and under a 12‐h photoperiod of blue light. Isolates within a given tetrad could be attributed to one of two sexual compatibility groups, on the basis of combinations of isolates leading to fertile crosses. Within each tetrad, mating type segregated in a 1:1 ratio, suggesting the existence of one mating type gene (MAT1) with two alleles MAT1–1 and MAT1–2. Under these same conditions, each individual isolate from an A‐group ascus originating from Brassica juncea produced pseudothecia with one or other of two compatible A‐group testers originating from B. napus. However, attempts to mate these eight A‐group single‐ascospore isolates with two B‐group single‐ascospore isolates of opposite mating types remained unsuccessful. This work is the first successful report of in vitro sexual reproduction between B‐group isolates, and provides evidence for bipolar heterothallism in B‐group isolates of L. maculans. The implications of this mating protocol for genetic studies of B‐group isolates and the consequences of this work for the taxonomy of L. maculans are discussed.
Spatial aggregation of pathotypes of barley powdery mildew
Authors: O'HARA R. B. , BROWN J. K. M.
Published: November 2003

Aggregation in the distribution of pathotypes of Erysiphe graminis f.sp. hordei, the barley powdery mildew pathogen, was investigated in field plots of ‘Golden Promise’, ‘Proctor’ and ‘Tyra’. ‘Golden Promise’ and ‘Proctor’ have no effective mildew resistance alleles, whereas ‘Tyra’ has Mla1, which was only effective against a proportion of the mildew population. Isolates of mildew were sampled according to a grid sampling scheme and their virulence spectra ascertained in order to group them according to pathotype. The populations were very diverse, and evidence for aggregation (quantified using join counts) was found only in the ‘Tyra’ plots, at distances of up to 1m. This aggregation was reduced in a subsequent sample. The results are consistent with a model in which mildew epidemics are started by a large number of initial infections, which then form diffuse, overlapping aggregations of clones. These aggregations then become more diffuse, so that the amount of aggregation reduces with time. The greater amount of aggregation seen in the ‘Tyra’ plots might have been caused by there being less initial inoculum with virulence towards that cultivar.
Leaflet abscission and phytoalexin production during the response of two faba bean breeding lines to Botrytis infection
Authors: HASHIM M. , ROBERTS J. A. , ROSSALL S. , DICKINSON M. J.
Published: November 2003

Comparisons were made between two breeding lines of faba bean (designated 224 and 335), following inoculation of leaf material with Botrytis cinerea and the more aggressive B. fabae. Line 335 exhibited greater initial resistance to infection, in terms of retarded lesion development and higher rates of phytoalexin production. However, in whole plant inoculation experiments line 335 abscinded its leaflets more readily after infection. The role of the gaseous plant hormone ethylene in this reaction was analysed. Both lines exhibited a similar abscission response to exogenous ethylene, applied at a concentration of 10 μL L−1. However, there were significant differences in the ability of the lines to produce ethylene. In response to inoculation with B. fabae, line 335 liberated ethylene at a higher concentration and for a longer period. These results suggest that the differential levels of abscission are a consequence of differences in the biosynthesis of ethylene in the two lines rather than in their responses to ethylene.
Transfer from Erwinia herbicola to Escherichia coli of a plasmid associated with biocontrol of fire blight
Authors: EL‐MASRY M. H. , BROWN T. A. , EPTON H. A. S. , SIGEE D. C.
Published: November 2003

Strains of Erwinia herbicola effective in the biocontrol of fire blight of hawthorn were used to investigate the possibility that the antagonistic activity is coded by plasmid‐born genes. Agarose gel electrophoresis of isolated plasmids from four antagonistic Erw. herbicola strains showed a band of a supercoiled 12 kb plasmid in each strain, with a second band greater than 16.2 kb consistently seen in two strains. Erw. herbicola strains showed resistance to penicillin‐G, which could be conferred on penicillin‐G sensitive Escherichia coli TG1 by transformation with a pure Erw. herbicola plasmid preparation. Transformed strains of Esc. coli appeared to contain the Erw. herbicola 12 kb plasmid, but not the > 16.2 kb plasmid. In an agar plate assay, Esc. coli transformants produced an inhibition zone against Erw. amylovora similar to those produced by the original Erw. herbicola strains. In two biocontrol assays, the transformed Esc. coli strains had a suppressive effect on disease development on infected pear fruit slices and hawthorn blossoms.
Zucchini yellow mosaic virus
Authors: DESBIEZ C. , LECOQ H.
Published: November 2003

Zucchini yellow mosaic potyvirus (ZYMV), first isolated in Italy in 1973, described in 1981, and then identified in all continents within a decade, is one of the most economically important viruses of cucurbit crops. It is efficiently aphid‐transmitted in a nonpersistent manner and it is also seed‐borne in zucchini squash, which could have contributed to its rapid spread worldwide. Biological variability has been observed among ZYMV isolates, concerning host range, symptomatology and aphid transmissibility. More recent studies also revealed a serological and molecular variability. The survival of ZYMV in areas where cucurbits are not grown throughout the year remains to be elucidated, because very few natural over‐wintering hosts have been identified so far. Partial control of ZYMV can be achieved by limiting transmission of the virus to the crops by aphids, using adapted cultural practices. Cross‐protection with a mild strain has been shown to be effective against most ZYMV isolates. Resistance genes found in cucurbit germplasms are currently being introduced into cultivars with good agronomical characteristics. Pathogen‐derived resistance strategies using the expression of ZYMV genes in transgenic plants have also been developed and appear promising. Nevertheless, the high biological variability of ZYMV justifies a careful evaluation of the deployment of genetic control strategies in order to increase their durability.
Screening Carica papaya × C. cauliflora hybrids for resistance to papaya ringspot virus‐type P
Authors: MAGDALITA P. M. , PERSLEY D. M. , GODWIN I. D. , DREW R. A. , ADKINS S. W.
Published: November 2003

Carica papaya, C. cauliflora and interspecific hybrids of these species were screened for resistance to two Australian isolates (338, 445) of papaya ringspot virus‐type P (PRSV‐P). Plants were manually inoculated with PRSV‐P in the glasshouse and the reaction assessed 30 days later by back‐inoculation to susceptible Cucurbita pepo and by a plate‐trapped antigen‐enzyme linked immunosorbent assay (PTA‐ELISA). Both parents and interspecific hybrids were also planted adjacent to infected C. papaya and 30 days later tested for PRSV‐P by PTA‐ELISA. All interspecific hybrid and C. cauliflora plants manually inoculated in the glasshouse or planted in the field failed to become infected, whereas C. papaya plants, in both situations, were infected by PRSV‐P. In addition, the surviving interspecific hybrid and C. cauliflora plants tested negative, while all C. papaya plants were positive for PRSV‐P in both the back‐inoculation and PTA‐ELISA tests. Thus, the interspecific hybrid and C. cauliflora plants were resistant to the Australian PRSV‐P isolates.
Genetic diversity of Fusarium moniliforme detected by vegetative compatibility groups and random amplified polymorphic DNA markers
Authors: HUANG R. , GALPERIN M. , LEVY Y. , PERL‐TREVES R.
Published: November 2003

Genetic diversity among Fusarium moniliforme isolates was analysed using vegetative compatibility group (VCG) and random amplified polymorphic DNA (RAPD) techniques. In the first experiment, RAPD was used to analyse a set of 43 isolates collected from different corn growing areas in Israel and the US. The isolates were assigned to 27 different VCGs. Thirty‐two RAPD haplotypes were also detected by analysing 48 polymorphic bands. RAPD could differentiate all the VCGs, except in two cases where two VCGs were assigned a single RAPD haplotype. In six cases, however, molecular variation was detected among isolates belonging to the same VCG. Cluster analysis of the RAPD data showed a very good agreement with the VCG grouping, e.g. isolates of the same VCG were always closely clustered by the molecular data. In a second experiment, 63 isolates of Fusarium moniliforme were collected from six corn lines growing in a single corn field. Extensive genetic variation was observed among the isolates: 42 different VCGs and 37 RAPD haplotypes were identified. Once again, RAPD patterns could differentiate nearly all the VCGs. However, in four cases, two different VCGs were grouped into a single RAPD haplotype, while in another three cases, isolates of the same VCG could be differentiated by distinct molecular haplotypes. The variation data was used to gain insight on the population structure and the patterns of genetic variation among geographical locations and within a single field. Hierarchical gene diversity analysis of the RAPD data indicated that most of the genetic variability (81%) was distributed within corn lines in the same field, suggesting that RAPD haplotype, or VCG frequencies, were not significantly affected by the plant genotypes grown in this experiment. Most of the RAPD band combinations did not display significant gametic phase disequilibrium, suggesting that active recombination might be occurring in the field. Our results indicate that by using a small number of primers, similar resolution was obtained by RAPD and VCG analysis, respectively. RAPD analysis is however, simpler to perform and its sensitivity in genotyping individuals within Fusarium moniliforme can be increased by analysing more primers, enabling a more detailed population genetic analysis of this important pathogen.
Presynaptic Membrane Receptor‐Reactive T Lymphocytes in Myasthenia Gravis
Authors: YI Q. , PIRSKANEN R. , LEFVERT A. K.
Published: November 2003

The majority of patients with myasthenia gravis were shown to have T and Bcells specific for a β‐bungarotoxin binding protein, presynaptic membranereceptor (PsmR). Such autoreactive T cells may be subdivided into differentsubsets according to the pattern of cytokine production. In this study theauthors examined the subpopulation of the T cells by analysing their IFN‐γand/or IL‐4 secretion pattern. T cell response to human muscle acetylcholinereceptor (AChR) was examined in parallel. PsmR‐stimulated IFN‐γ secretionwas found in 60%, and IL‐4 secretion in 48% of the patients. Cells stimulated tosecrete both IFN‐γ and IL‐4 or IFN‐γ only were the most common patterns.Treatment of the cells with a mouse anti‐human HLA‐DR antibody abolished thesecretion of cytokines. There was a positive correlation between the numbers ofPsmR‐reactive and AChR‐reactive T cells. In conclusion, the results show thatPsmR‐stimulated T cells secreted IFN‐γ and/or IL‐4. This T cell response isMHC class II restricted. Thus, this study indicates that both Th1/Th2 or Th0subsets of the T cells are involved in the autoimmune response in thedisease.
Limited T‐Cell Receptor V Gene Usage in Inclusion Body Myositis
Authors: FYHR I.‐M. , MOSLEMI A.‐R. , TARKOWSKI A. , LINDBERG C. , OLDFORS A.
Published: November 2003

Inclusion body myositis (IBM) is a chronic, progressive inflammatory myopathy. The inflammatory infiltrates are dominated by T cells, which frequently invade muscle fibres. The present study was performed to characterize the usage of the variable (V)segment of the T‐cell receptor of muscle infiltrating cells in IBM. Using the reverse transcriptase polymerase chain reaction (RT‐PCR) technique the authors analysed the expression of 22 Vα and 24 Vβ families in muscle tissue from six patientswith IBM displaying intense inflammatory cell infiltration. The following Vα/Vβ families appeared in at least 50% of the patients: Vα1, 5, 7, 15, 16, 17, 20, 21, 22 and Vβ3, 5.2, 8, 12, 14, 22. In all patients Vα7, 16 and Vβ8 wereexpressed in muscle tissue. Furthermore, in two of the patients peripheral blood lymphocytes (PBL) were investigated in parallel. There was a restricted usage of Vα and Vβ families in muscle in comparison to PBL, indicating a selective homing orlocal proliferation of T lymphocytes in the inflammatory lesions in IBM.
Fcγ Receptor Expression on Sheep Afferent Lymph Dendritic Cells and Rapid Modulation of Cell Surface Phenotype Following Fcγ Receptor Engagement In Vitro and In Vivo
Authors: COUGHLAN S. N. , HARKISS G. D. , DICKSON L. , HOPKINS J.
Published: November 2003

Afferent lymph dendritic cells were analysed for the presence of Fcγ receptors by Western blotting and for modulation of surface markers following Fcγ receptor engagement in vitro and in vivo. The results showed thatunstimulated dendritic cells expressed FcγRII constitutively. When dendritic cells were incubated in vitro with antigen/antibody complexes in antibody excess, a marked reduction in surface staining was observed for MHC class II, CD1, CD44,and VLA‐4 after 8 h in culture. These changes did not occur with antigen or antibody alone. DC expression of LFA‐1 and LFA‐3 were slightly reduced after 8 h in culture with Ova alone, but this was enhanced slightly when the cells werecultured with immune complexes. Even more marked reductions in surface staining for MHC class II, CD1, CD44 and VLA‐4 were observed on dendritic cells 4–8 h following secondary antigen challenge in vivo. LFA‐1 and LFA‐3 expressionwas reduced only slightly. The level of expression of MHC class II, CD1, LFA‐1 and LFA‐3 was substantially increased over resting values 24 h after FcγR occupancy. The intensity of staining at this time was also significantly elevated for CD44, LFA‐1, LFA‐3 and VLA‐4. These results show that engagement of Fcγ receptors cause a substantial modulation of the dendritic cell surface phenotype after immune complex uptake. The phenomenon may function to maximize subsequent presentation of thechallenge antigen to T cells.
Interleukin 10 Release During Endotoxaemia in Chimpanzees: Role of Platelet‐Activating Factor and Interleukin 6
Authors: VAN DER POLL T. , JANSEN J. , LEVI M. , TEN CATE H. , HACK C. E. , AARDEN L. A. , TEN CATE J. W. , VAN DEVENTER S. J. H.
Published: November 2003

Interleukin (IL‐)10 has been demonstrated to inhibit endotoxin‐induced production of a number of pro‐inflammatory cytokines. The present study sought to compare the appearances in the circulation of IL‐10, IL‐6 and IL‐8, and to assess the roles ofendogenously produced platelet‐activating factor (PAF) and IL‐6 in IL‐10 release during endotoxaemia in chimpanzees. Intravenous injection of endotoxin (lot EC‐5, 4 ng/kg, n = 8) induced a transient rise in serum IL‐10concentrations, peaking after 2 h (213 ± 70 pg/ml; P < 0.05). No correlations existed between peak IL‐10 levels, and peak IL‐6 and IL‐8 levels. Neither infusion of the specific PAF antagonist TCV‐309(n = 4), nor infusion of a neutralizing anti‐IL‐6 monoclonal antibody (n = 4) influenced endotoxin‐induced IL‐10 release. IL‐10 release elicited by injection of endotoxin is not mediated by PAF or IL6. Tom van der Poll MD, Academic Medical Center, Department of Internal Medicine, F4222, Meibergdreef 9, 1105 AZ Amsterdam, the Netherlands
Acquisition of CD40 Expression During Murine B‐Cell Differentiation
Authors: GRANDIEN A. , BRÁS A. , MARTINEZ‐A C.
Published: November 2003

Expression of CD40 on mouse cells was investigated comparing the binding to cells of a monoclonal antibody against CD40, to that of a soluble fusion protein consisting of the extracellular domains of the mouse CD40‐ligand. The analysis of a series ofestablished cell lines failed to demonstrate expression of CD40 on pro‐ or pre‐B cells, and indicated that CD40 expression was restricted to cells that had undergone productive heavy‐ and light‐chain gene rearrangements, and expressed surface Ig. In cells from normal mice, CD40 first becomes detectable, although at low levels, on a subset of small pre‐B‐II cells in bone marrow, the levels of CD40 expression increasing thereafter during B‐cell maturation. Thus, immature B cells (IgM+IgDlo B220lo) express intermediate levels of CD40, and mature B cells (IgM+ IgDhi B220hi) express high levels of CD40. Anatomical location also seems to correlate with the levels of CD40expression, as B cells expressing the highest levels of CD40 were found in lymph nodes and Peyer’s patches.
Oral Tolerization Leads to Active Suppression and Bystander Tolerance in Adult Rats while Anergy Dominates in Young Rats
Authors: LUNDIN B. S. , DAHLGREN U. I. H. , HANSON L. Å. , TELEMO E.
Published: November 2003

Oral tolerance was induced in 4‐week‐old (young) and 12‐week‐old (adult) rats by feeding ovalbumin (OvA)‐containing pellets during 4 weeks. Seven weeks after removal of the OvA‐pellets the rats were immunized with a mixture of OvA and human serumalbumin (HSA) in Freund’s complete adjuvant (FCA), and the following immune response was monitored. Both the young and adult groups of OvA‐fed rats had significantly suppressed OvA‐specific delayed‐type hypersensitivity (DTH) responses and T‐cellproliferation, reflecting a long‐lasting T‐cell tolerance to OvA both in vivo and in vitro. Furthermore, spleen cells from rats tolerized as adults were able to suppress the proliferation of primed T‐cells from normal immunized rats, demonstrating thepresence of antigen‐specific suppressive cells. Accordingly, the adult rats showed bystander suppression of the response to HSA with respect to DTH‐reaction, specific proliferation, and reduced enlargement of the draining lymph nodes after immunization.There was no evidence of active suppression in vitro or bystander tolerance in the orally tolerized young group, indicating that anergy rather than active suppression was prevalent in these rats. Furthermore, in the young group there was no suppression of the antibody response since the IgG and IgE anti‐OvA antibody levels were indistinguishable from those of the controls. Contrary to the young rats, the adult fed group showed transiently elevated levels of IgG anti‐OvA antibodies at 1 weekpost‐immunization, followed by a subsequent significantly suppressed IgG antibody response. In conclusion, the results demonstrate that the induction of anergy or active suppression after antigen feeding can be determined by the age at which the antigenis introduced to the mucosal immune system.
HLA‐B27‐Restricted Cytotoxic T Lymphocyte Responses to Arthritogenic Enterobacteria or Self‐Antigens are Dominated by Closely Related TCRBV Gene Segments. A Study in Patients with Reactive Arthritis
Authors: DUCHMANN R. , MAY E. , ACKERMANN B. , GOERGEN B. , MEYER ZUM BÜSCHENFELDE K.‐H. , MÄRKER‐HERMANN E.
Published: November 2003

Identification of the T‐cell receptors (TCR) used by synovial cytotoxic T lymphocytes (CTL) of patients with reactive arthritis (ReA) may be crucial to better understanding the pathogenetic mechanism underlying the HLA‐B27 association ofspondylarthropathies. The authors, therefore, sequenced 25 TCRB chains from HLA‐B27‐restricted CD8+ CTL clones and two clonal lines specific for self‐ or Yersinia enterocolitica antigen isolated from synovial fluids of 3HLA‐B27+ patients with ReA and PBL of one healthy HLA‐B27+ individual. Fourteen non‐HLA‐B27‐restricted CTL served as controls. Both autoreactive and Y. enterocolitica specific HLA‐B27‐restricted CTL used a highlylimited set of VB genes with preferential rearrangement of three closely related VB families (VB 13,14,17), suggesting that these families contain a preferred site for contact with the HLA‐B27 molecule. In addition, the presence of limited TCRBJ usage,limited heterogeneity in CDR3 sequences and dominant clones from individual donors among these CTL indicate that TCRB chain usage is further restricted by a limited set of peptides bound to the HLA‐B27 molecule. Limited TCR usage by SF CTL of ReA patients may lay a basis for therapeutical manipulation of the T‐cell response in the spondylarthropathies.
Gliadin Specific, HLA DQ2‐Restricted T Cells are Commonly Found in Small Intestinal Biopsies from Coeliac Disease Patients, but not from Controls
Authors: MOLBERG Ø . , KETT K. , SCOTT H. , THORSBY E. , SOLLID L. M. , LUNDIN K. E. A.
Published: November 2003

The authors have analysed gliadin specific, CD4+ T cells isolated from small intestinal biopsies of 23 adult coeliac disease patients (20 on a gluten‐free diet and three untreated) and nine control patients. The biopsies were stimulated ex vivo with a peptic/tryptic digest of gliadin for 24 h, and activated T cells were positively selected with paramagnetic beads coated with an antibody against the interleukin‐2 receptor. The T cells were expanded and tested for gliadin reactivity and HLA restriction. Gliadin specific, polyclonal T cell lines were recovered from biopsies of all 23 patients. Inhibition studies of T cell lines from 21 patients with anti‐HLA monoclonal antibodies indicated predominant presentation of the gliadin antigen by HLA‐DQ2 in T cell lines from 11 patients (lines from seven patients with complete MoAb inhibition, the remaining with incomplete inhibition) and incomplete inhibition by HLA‐DR3 in lines from three patients. Nine gliadin specific T cell clones from six patients were established; all of these were HLA‐DQ2 restricted. Gliadin specific T cells were not found in biopsies from the non‐coeliac controls. Our findings demonstrate that gliadin reactive T cells are commonly found in the intestinal mucosa of CD patients and they support the notion that the majority of T cells recognize gliadin peptide(s) when presented by the disease associated DQ2 molecules.
Serum Antibodies from Patients with Primary Sjögren’s Syndrome and Systemic Lupus Erythematosus Recognize Multiple Epitopes on the La(SS‐B) Autoantigen Resembling Viral Protein Sequences
Authors: HAAHEIM L. R. , HALSE A.‐K. , KVAKESTAD R. , STERN B. , NORMANN Ø. , JONSSON R.
Published: November 2003

The aim of this study was to investigate the epitope recognition pattern of La(SS‐B) autoantibodies in sera from patients with Sjögren’s syndrome (SS) and systemic lupus erythematosus (SLE) using overlapping synthetic decapeptides on solidphase. Eighty different decapeptides with five amino acids overlap from the human La(SS‐B) autoantigen were synthesized on cellulose paper using F‐moc chemistry. Tests were performed with 14 SS and six SLE sera. The results showed that the immune response to the La(SS‐B) oligopeptides was restricted and unique for each individual with no particular pattern typical for each of the two diseases, apart from the fact that SLE sera gave positive reaction with fewer peptides. Regions within the N‐ and C‐termini harboured most of the positive sequences. The authors specifically addressed the possibility of a viral aetiology for disease development or autoantibody generation. In this context the most frequently recognized linear epitopes on the La(SS‐B)autoantigen showed sequence similarities with proteins from a range of ubiquitous human viruses, in particular from the herpes virus group. The La(SS‐B) autoantibodies may thus be generated through molecular mimicry.
Expression and Characterization of a Divalent Chimeric Anti‐Human CD3 Single Chain Antibody
Authors: MA SHENGLIN , THOMPSON J. , HU HUAIZHONG , NEVILLE JR D. M.
Published: November 2003

Murine anti‐CD3 monoclonal antibodies (MoAbs) are used in clinical practice for immunosuppression. However, there are two major drawbacks to this treatment: the associated cytokine release syndrome and human anti‐mouse antibody response. To overcome these side‐effects, the authors generated a chimeric anti‐human CD3 single chain antibody, scUCHT1. It is an IgM variant of UCHT1, a mouse IgG1 MoAb directed against human CD3. scUCHT1 consists of the light and heavy variable chain binding domains of UCHT1 and a human IgM Fc region (CH2 to CH4). scUCHT1 was produced by COS‐7 and SP2/0 transfectants, and mainly assembled in a dimeric form. It retained the binding specificity and affinity of the parental MoAb UCHT1. In contrast to UCHT1, scUCHT1 did not induce T‐cell proliferation and cytokine release (TNF‐α and IFN‐γ) in in vitro assays. These results suggest that the engineered chimeric anti‐CD3 single chain antibody (scUCHT1) may be useful in clinical immunosuppressive treatment.
Sharing of Four DR‐β Sequence Motifs Between HLA‐DRB1*1601 and DRB1*1101 Correlates with Frequent Degenerate T‐Cell Recognition of HA306–320 Peptide Complexed to these Two Molecules
Authors: ZELISZEWSKI D. , DORVAL I. , GOLVANO J. J. , PREVOST A. , BORRAS‐CUESTA F. , STERKERS G.
Published: November 2003

This paper shows that the seven HA306–320 specific T‐cell clones isolated from one individual recognize the peptide complexed to both autologous HLA–DRB1*1101 and allogeneic HLA‐DRB1*1601 (or DRB5*0201) molecules. For each T‐cell clone, a single T‐cell receptor (TCR) is involved in the recognition of these two different peptide‐DR complexes as evidenced by cold target competition experiments. Yet, the seven T‐cell clones express several different TCRs as judged by Vβ‐Jβ usage and fine specificities. Furthermore, one representative clone has the same fine specificity for HA306–320 analogues mutated at epitopic residues irrespective of the use of DR1101 or DR1601 APC. These results suggest that structural differences between DRB1*1101 and DRB1*1601 (or DRB5*0201) do not dramatically influence the orientation of HA306–320 in the grooves such that most residues interacting with TCRs are conserved. In another individual, the same pattern of restriction, i.e. DR1101 + DR1601, was found for several HA306–320 specific clones. Two additional patterns, DR1101 + DR0801 and DR1101 + DR0801 + DR1601, were identified. By comparing DR sequences the authors found that DRB1*1101 and DRB1*1601 share four important motifs, i.e. β85–86, β67–71, β57 and β28–31 supposed to line three distinct HLA‐DR pockets. Three of these motifs are also shared with DRB1*0801. All the results further support that the motif similarities allow the peptide to adopt very similar orientations in the cross‐reacting DR molecules.
Experimental Human Plasmodium Falciparum Infections: Longitudinal Analysis of Lymphocyte Responses with Particular Reference to γδ T Cells
Authors: RZEPCZYK C. M. , STAMATIOU S. , ANDERSON K. , STOWERS A. , CHENG Q. , SAUL A. , ALLWORTH A. , McCORMACK J. , WHITBY M. , OLIVE C. , LAWRENCE G.
Published: November 2003

The kinetics of the γδ T‐cell response was analysed in the context of the overall haematological response in subjects experimentally infected with sporozoites of Plasmodium falciparum. Numbers of γδ and αβ T cells and NK cells declined markedly during infection to reach minimum values 12–13 days post‐infection when the patients were ill. This decline commenced from the beginning of the erythrocytic cycle and well before parasites could be detected microscopically and clinical symptoms developed. Platelet numbers also declined. In vivo activation of γδ T cells was evident with sequential up‐regulation of the activation markers CD69 and HLA‐DR. γδ T cell numbers were highest after treatment with the majority being CD4−CD8−, HLA‐DR+ and showing reduced CD45RA expression. Contrary to some published observations γδ T‐cell percentages remained within the normal range. Little evidence of up‐regulation of activation or memory markers was observed in the αβ T‐cell population. In vitro proliferative responses to malaria antigen which involve γδ T cells were lost as the infection progressed and the lymphocyte count declined but these could be restored with the addition of exogenous IL‐2 to cultures. The authors findings are consistent with a protective and/or immunomodulatory role for γδ T cells in malaria.
Role of Monocytes in the Up‐Regulation of the Early Activation Marker CD69 on B and T Murine Lymphocytes Induced by Microbial Mitogens
Authors: VILANOVA M. , TAVARES D. , FERREIRA P. , OLIVEIRA L. , NÓBREGA A. , APPELBERG R. , ARALA‐CHAVES M.
Published: November 2003

CD69 is an early marker of lymphoid cell activation. The authors report on an up‐regulation of CD69 in splenic B and T cells of C57Bl/6 mice after administration of lipopolysaccharide (LPS) or microbial immunosuppressive/mitogenic (ISM) proteins produced by C. albicans (p43) and African Swine Fever Virus (p36). This up‐regulation of CD69 was observed 6 and 24 h after mitogenic treatments. The same pattern of increased CD69 expression was observed in the lymph nodes of mice treated with p43 or LPS, whereas p36 treatment failed to induce increased CD69 expression in this organ. Intracellular calcium mobilization was induced in splenic B and T lymphocytes after incubation of total spleen cells with LPS, p43 or p36. This increase was higher in B than in T cells. Increased calcium mobilization was also seen in lymph node B cells after incubation with p43 or p36 and in lymph node T cells after p43 stimulation. Up‐regulation of CD69 expression on B and T cells was also observed after in vitro stimulation of spleen cells with the three mitogens used. Similar results were obtained with culture supernatants of macrophage/monocyte (Mφ) cells activated with LPS (LPS/MφCS). Stimulation of Mφ cells with LPS or with the ISM proteins is demonstrated by the increased production of nitrites by these cells. The increased in vitro expression of CD69 was, however, not abolished by monoclonal antibodies to Mφ cytokines such as IL‐6, IL‐10 or TNFα. No increased expression of CD69 was found in vitro on purified B or T cells, even when mixed upon stimulation with p43, p36, LPS or with LPS/MφCS. However, an increase in the expression of CD69 was observed on B cells co‐cultured with Mφ cells after treatment with LPS or p36. All three mitogens failed to induce increased CD69 expression on cultured T cells mixed with Mφ cells.
Down‐Regulation of Anti‐CD3 Antibody‐Induced IL‐4 Production by Bovine Caseins Hydrolysed with Lactobacillus GG‐Derived Enzymes
Authors: SÜTAS Y. , HURME M. , ISOLAURI E.
Published: November 2003

A prerequisite for systemic hyporesponsiveness to dietary antigens is their processing in the gut. This study investigated whether bovine caseins degraded by enzymes of an intestinal bacterial strain, Lactobacillus GG (ATCC 53103), could regulate the cytokine production by anti‐CD3 antibody‐induced peripheral blood mononuclear cells of 14 atopic patients, aged 5–29 (mean, 16) months. Purified casein up‐regulated the interleukin‐4 and interferon‐γ production, P = 0.008 and P = 0.008, respectively. Conversely, Lactobacillus GG‐degraded casein down‐regulated the interleukin‐4 production, P = 0.003, with no effect on interferon‐γ. These results indicate that intestinal bacteria may modify immunomodulatory properties of native food proteins and introduce a promising tool to provide protection from potentially harmful dietary antigens at a young age.
Murine Acariasis. II. Immunological Dysfunction and Evidence for Chronic Activation of Th‐2 Lymphocytes
Authors: JUNGMANN P. , FREITAS A. , BANDEIRA A. , NOBREGA A. , COUTINHO A. , MARCOS M.‐A. , MINOPRIO P.
Published: November 2003

The authors describe the immunological profile of BALB/c mice with Mite‐Associated Ulcerative Dermatitis (MAUD)‐like disease, due to Myocoptes musculinus (Koch 1844) infestation. The disease probably involves allergic mechanisms and is characterized by erythematous and pruritic skin lesions, widespread hair loss, lymphadenopathy, lymphocytopenia, granulocytosis and wasting. Affected individuals had much reduced numbers of pre‐B and B cells in bone marrow and B cells in blood; decreased T‐cell numbers in peripheral lymphoid organs and blood; hypergammaglobulinaemia with selective increases of IgG1, IgE and IgA, and depletion on IgM and IgG3, the same isotype distribution being detected in splenic plasmocytes; qualitative modifications of the serum antibody reactivity pattern; and increased production of IL‐4 with decreased IL‐2 production after in vitro polyclonal stimulation of T cells. Taken together, these results suggest that infestation by M. musculinus in BALB/c mice leads to a significant immunological disorder resulting in a T‐helper‐2 (Th‐2) type response, with marked systemic consequences. This pathological condition may thus provide a useful model system for the immunobiological perturbation associated with chronic allergic disease.
Both Murine SAA1 and SAA2 Yield AA Amyloid in Alveolar Hydatid Cyst‐Infected Mice
Authors: BELL A. W. , CHAN S. L. , MARCANTONIO D. , ALI‐KHAN Z.
Published: November 2003

Amyloid susceptible C57BL/6 and partially amyloid resistant A/J mice, infected intraperitoneally with 250 alveolar hydatid cyst (AHC), the larval stage of a cestode parasite Echinococcus multilocularis, develop multiple organ amyloid deposits at approximately 1 and 4 weeks post infection (p.i.), respectively. Pooled spleens and livers from each mouse strain, at 8 and 10 weeks p.i., were used for the purification of protein AA utilizing a HiLoad Superdex 200 column equilibrated with 5 M guanidine‐HCl. Protein AA from each mouse strain was separated on 16% Tris‐tricine SDS‐PAGE gels and immunoblotted with monospecific rabbit anti‐mouse AA IgG; five and six immunoreactive AA subspecies were detected in the C57BL/6 and A/J materials, respectively. N‐Terminal amino acid sequence analysis was performed on the bulk column‐purified protein AA as well as on the electroblotted AA subspecies from each mouse strain. The results show a mixture of serum amyloid A1 (SAA1) and (SAA2)‐derived AA protein from each mouse strain; SAA1‐derived AA, although alluded to, has never been demonstrated as tissue deposits in mice. These findings suggest that the intense and persistent inflammatory processes in AHC‐infected mice may have induced conversion of weakly amyloidogenic SAA1 to AA. This conversion could be detected by amino acid sequencing of electrophoretically separated AA subspecies.
Human T‐Cell Epitopes on the Mycobacterium tuberculosis Secreted Protein MPT64
Authors: ROCHE P. W. , FENG C. G. , BRITTON W. J.
Published: November 2003

Mycobacterium tuberculosis secretes a number of proteins into the extracellular envirornment, some of which are restricted to the M. tuberculosis complex. These proteins are targets for T‐ and B‐cell immune responses in tuberculosis (TB) patients and their contacts. The authors have mapped the immunogenic regions of the MPT64 protein of M. tuberculosis using peripheral blood mononuclear cells (PBMC) from TB patients and a set of overlapping peptides encompassing the complete sequence of the protein. T‐cell epitopes which induced proliferation or interferon‐gamma (IFN‐γ) release were distributed over the full length of the protein. A C‐terminal region of the protein, however, contains sequences recognized in the context of multiple HLA‐DR phenotypes by more than 80% of the subjects tested. The nature of the T‐cell response was further investigated by generating MPT64‐specific T‐cell lines. These lines also identified the T‐cell epitopes in the C‐terminal region of the protein. On stimulation with antigen the lines secreted IFN‐γ, but not interleukin 4 (IL‐4). A minority of TB patients (6/32) mounted an antibody response to MPT64. Sera from half (3/6) of these identified two linear antibody binding sites. These results confirm the significance of this protein in the immune response to tuberculosis infection.
Indirect Effects are Independent of the Way of Tolerance Induction
Authors: CARVALHO C. R. , VAZ N. M.
Published: November 2003

Oral tolerance is a T‐cell mediated phenomenon defined by a refractoriness to parenteral immunization with a protein that was first contacted by oral route. However, the authors have shown that the injection of a tolerated protein is not neutral for the immune system. In mice made tolerant to KLH by gavage, co‐immunization with KLH and DNP‐Ova blocks anti‐DNP antibody formation. Anti‐DNP antibody formation resulting from immunization with DNP‐Ova can also be blocked by co‐immunization with a dietary protein (zein) or a self component (fibrinogen). The inhibitory effects resulting from immunization with a tolerated protein, designated indirect effects, do not affect the induction of oral tolerance to another protein. These results support the hypothesis that active mechanism are involved in the maintenance of tolerance.
Antibodies Reactive to Trypanosoma cruzi Epimastigotes or Amastigotes Express Different Idiotypic Patterns if from Patients with Different Clinical Forms of Chagas' Disease
Authors: RIBEIRO‐RODRIGUES R. , COLLEY D. G. , CORREA‐OLIVEIRA R. , CARTER C. E.
Published: November 2003

Antibodies (Abs) were purified from pooled sera of patients with either indeterminate (IND or I) or cardiac (CARD or C) Chagas' disease, on either epimastigote (EPI or E) or amastigote‐enriched (AMAST or A) antigen (Ag) columns and their idiotypic (Id) expression examined. Anti‐Id rabbit Abs were raised to the different preparations (E‐IdI, E‐IdC, A‐IdI and A‐IdC). Competitive ELISAs using anti‐Ids were able to discriminate between IdI and IdC, disregarding Ag reactivity. E‐IdI and A‐IdI present different inhibitory abilities, as do E‐IdC and A‐IdC, but IdC always competes with IdI for anti‐IdI comparably. In contrast, a 4–8‐fold increase of IdI is required to compete in parallel with IdC for anti‐IdC. Therefore, Ids from IND patients share only low levels of the Ids that are most characteristic of CARD patients. While some CARD Abs also express Ids in common with IND patients, these studies reveal that CARD Abs express some Ids that are characteristic to only CARD patients, and these Ids are present on Abs purified with either EPI or AMAST.
Heterogeneous Expression of Recombination Activating Genes and Surface CD5 in CD3low CD4+CD8+ Thymocytes
Authors: SHEARD M. , SUGAYA K. , FURUTA M. , TSUDA S. , TAKAHAMA Y.
Published: November 2003

Clonal selection in the thymus occurs mostly in the CD4+CD8+ (double positive; DP) stage. Within DP thymocytes, cells in the CD3low subset are believed to be involved in clonal selection, and this subset is generally considered as a homogeneous population. T‐cell antigen‐receptor (TCR) signals on DP thymocytes are known to (i) down‐regulate recombination activating gene (RAG) expression; and (ii) up‐regulate the expression of T‐cell function‐associated molecules, including the cell surface glycoprotein CD5. The present study examined the expression of RAG and CD5 molecules among the subpopulations of normal adult DP thymocytes. DP thymocytes were fractionated according to their cell surface expression levels of TCR‐CD3 complex into CD3dim, CD3lo, CD3med, and CD3hi cells, since TCR expression is known to increase during thymocyte maturation. Down‐regulation of RAG mRNA was located between the CD3low and CD3med DP populations. However, within the DP CD3low subpopulation, we found that CD5 varies from low to high expression levels. Upon fractionation of DP CD3low thymocytes into CD5low and CD5high subsets, we were able to detect down‐regulation of RAG transcripts within the CD3low subpopulation. Thus, by the criteria of CD5 surface expression and RAG mRNA expression levels, DP CD3low thymocytes can be considered a heterogeneous thymocyte subpopulation.
Immunophenotyping of Human Bone Marrow‐Derived Macrophages
Authors: KÖLLER M. , WILLHEIM M. , KRUGLUGER W. , KURZ M. , HÖCKER P. , FÖRSTER O. , BOLTZ‐NITULESCU G.
Published: November 2003

In vitro cultured cells derived from human bone marrow stimulated with macrophage‐colony stimulating factor (M‐CSF) or granulocyte‐macrophage‐colony stimulating factor (GM‐CSF) were investigated for their immunophenotypic surface characteristics by flow cytometry. Approximately 80–90% of the cells showed morphological and histochemical features of macrophages and bore CD11a, CD11b, CD11c, CD14, CD29, CD32, CD33, CD44, CD45, CD54, CD64, CD71, HLA‐DR antigen. The expression of CD4, CD25 and CD45RO were detected only in low density. Bone marrow‐derived macrophages were negative for CD8, CD45RA, CD16 and CD23. Functionally, macrophages were able to phagocytose opsonized Escherichia coli, but no oxidative bursts were induced either by fmlp or by opsonized bacteria. Cell surface phenotyping revealed a CD pattern very similar to monocyte‐derived phagocytes, and was partially distinct from resident stromal macrophages.
Fibronectin Co‐Stimulates via the α5β1 Receptor IL‐2, IL‐4 Production by Splenic, Granuloma Lymphocytes of Schistosoma mansoni Infected Mice
Authors: ZHU Y. , BOROS D. L.
Published: November 2003

In murine Schistosomiasis mansoni, soluble worm egg antigens (SEA) induce L3T4+ T helper cell‐mediated chronic granulomatous inflammations around parasite eggs. Within the fully developed granuloma lymphocytes, macrophages, and eosinophils, fibroblasts are embedded in extracellular matrix (ECM) composed of fibronectin, laminin, glycosaminoglycans and collagens. The present study examined in vitro the putative co‐stimulatory role of fibronectin (FN) in acute and chronic infection splenic and granuloma lymphocyte responses. Plate‐bound FN enhanced the anti‐CD3 MoAb stimulated normal and acute or chronic infection splenic lymphoproliferation by 20–32%. The co‐stimulatory effect was evident in SEA stimulated acute but not chronic infection spleen cells. Proliferation of stimulated granuloma lymphocytes could not be up‐regulated by immobilized FN. Plate‐bound FN significantly enhanced IL‐2 and IL‐4 production by SEA‐stimulated acute, but not chronic, infection granuloma lymphocytes. However, FN had no influence on the high level of IL‐2, IL‐4 production of anti‐CD3 MoAb stimulated acute or chronic infection splenic or granuloma lymphocytes. Because in the antigen‐stimulated acute infection spleen or granuloma cultures the co‐stimulatory effect by FN was abrogated by the tripeptide (RGD) arg‐gly asp, and anti α5β1 antibody, enhancement is attributed to signalling via the α5β1 integrin receptor of lymphocytes.
Co‐Stimulation and Co‐Inhibition: Equal Partners in Regulation
Authors: SINCLAIR N. R. StC. , ANDERSON C. C.
Published: November 2003

Specific immune responses are controlled by two counterbalancing mechanisms—co‐stimulation and co‐inhibition. Antigen receptors determine specificity, activate co‐stimulation and/or co‐inhibition, and interact with these co‐stimulatory/co‐inhibitory mechanisms to dictate the direction of the immune response, either positive or negative. Co‐stimulatory or co‐inhibitory ligands interact with their specific receptors and may indicate the context in which antigen is perceived by lymphocytes. Ligation of antigen receptors may activate only co‐stimulatory or co‐inhibitory mechanisms, and thus may influence secondarily the direction of the immune response. Furthermore, the activity of a given co‐stimulator or co‐inhibitory receptor is modified depending on signalling via the antigen receptor. If neither co‐stimulators nor co‐inhibitors are present, lymphocytes, activated in response to antigen receptor signalling, produce low levels of effector elements and then revert to inactivity. Co‐inhibitors are defective in autoimmune disease.
Activation of the Protein Kinase A Increases the DNA‐Binding and Transcriptional Activity of c‐Rel in T Cells
Authors: LAHDENPOHJA N. , HENTTINEN T. , HURME M.
Published: November 2003

Cyclic AMP (cAMP)‐dependent protein kinase A (PKA) is known to have both negative and positive effects on the activation mechanisms of T lymphocytes. The authors have analysed the effect of increased cAMP on the activation of NF‐κB transcription factor. This factor controls the expression of several genes (e.g. IL‐2 and IL‐2 receptor) involved in the activation and proliferation of T cells. The authors found that elevation of intracellular cAMP in Jurkat T leukaemia cells activated with phorbol ester (PDBu)/calcium ionophore (A23187) increased the DNA‐binding of NF‐κB as detected by the electrophoretic mobility shift assay (EMSA). Analysis of the subunit composition of the DNA‐binding complex indicated that the amount of c‐Rel was enhanced while RelA was decreased. Analysis of the effect of elevated cAMP on the degradation of IκB‐α and IκB‐β did not reveal an essential change in degradation kinetics of these inhibitor proteins. The elevation of cAMP did not increase the synthesis of c‐Rel, but it enhanced the nuclear localization of this protein. Transfection of Jurkat cells with a plasmid kB/TK10‐CAT indicated that the increased DNA‐binding of c‐Rel containing complexes seen in EMSA was also functional. These data imply that the strong and long‐lasting c‐Rel nuclear localization and DNA‐binding induced by protein kinase A is not due to increased c‐Rel synthesis or enhanced degradation of the IκB inhibitors. Therefore, a direct phosphorylation of the c‐Rel protein is the most plausible explanation for these observations. Taken together, these results suggest that cAMP is able to regulate the expression of NF‐κB‐dependent genes in T cells by modifying the composition and subunit activity of NF‐κB.
Serum Level of Interleukin‐4 in Patients with Perennial Allergic Rhinitis During Allergen‐Specific Immunotherapy
Authors: OHASHI Y. , NAKAI Y. , OKAMOTO H. , OHNO Y. , SAKAMOTO H. , SUGIURA Y. , KAKINOKI Y. , TANAKA A. , KISHIMOTO K. , WASHIO Y. , HAYASHI M.
Published: November 2003

Interleukin‐4 (IL‐4) may play a central role in the IgE synthesis system, the development of Th‐2‐like cells, and co‐ordination as well as the persistence of airway inflammatory process in allergic disorders. Therefore, IL‐4 plays a key role in airway allergic disorders. This study aimed at investigating the serum concentrations of IL‐4 in patients with perennial allergic rhinitis, with special reference to the possible changes and the clinical relevance following long‐term immunotherapy. The study has demonstrated that the serum level of IL‐4 in allergic rhinitis patients before immunotherapy is significantly higher than that in non‐atopic individuals. However, the serum IL‐4 level in allergic rhinitis patients did not decrease following anti‐allergic medications but significantly decreased following immunotherapy. The percentage decrease in IL‐4 was correlated significantly with the percentage decrease in specific IgE antibodies following long‐term immunotherapy. Immunotherapy also significantly decreased specific IgE antibodies, but this reduction in specific IgE antibodies was not significantly correlated with the clinical improvement. In contrast, the percentage decrease in serum IL‐4 was significantly correlated with the percentage decrease in symptomatic scores. The authors interpret these data to mean that immunotherapy alters T‐cell cytokine profiles in the long‐term, and a decline of IL‐4 following immunotherapy could modulate not only production of specific IgE antibodies but also inflammatory cellular events, leading to symptomatic relief in allergic rhinitis.