bluuebunny/crossref_metadata_2025 huggingface dataset:
Study of antigenic sites on the asialoglycoprotein receptor recognized by autoantibodies
Authors: Hajoui O , Martin S , Alvarez F
Published: March 2003

The aim of this study was to identify the epitopes recognized by antibodies to the asialoglycoprotein receptor, a specific hepatocyte protein, from sera of patients with autoimmune hepatitis. An ELISA test was used to detect anti-asialoglycoprotein receptor antibodies in the sera of patients with autoimmune hepatitis. Positive sera were tested against the same antigen by slot blot, by Western blot and by immunoprecipitation of the untreated protein and following treatment with β-mercaptoethanol (β-ME) and endoglycosidase F. The mature, unglycosylated and partially glycosylated forms of the asialoglycoprotein receptor synthesized by HepG2 cells were tested against positive patients' sera, as well as the in vitro translated unglycosylated form of the H1 subunit of the receptor. Sera from patients with autoimmune hepatitis recognized equally the native form, as well as the β-ME-modified form, but less well the deglycosylated form of the human mature receptor. No reactivity was found when these sera were tested against the denatured human protein. In addition, neither the unglycosylated H1 subunit nor any of the HepG2-synthesized asialoglycoprotein receptor forms bound to the antibodies. Altogether, these results show that anti-asialoglycoprotein receptor antibodies in the sera of patients with autoimmune hepatitis are directed against conformational structures of the mature hetero-oligomeric form of the human liver protein and that at least some epitopes were located on the extracellular domain of the antigen.
Perioperative cytokine release during coronary artery bypass grafting in patients of different ages
Authors: Roth-Isigkeit A , Schwarzenberger J , v Borstel T , Gehring H , Ocklitz E , Wagner K , Schmucker P , Seyfarth M
Published: March 2003

Surgical interventions and cardiopulmonary bypass (CPB) induce a systemic inflammatory response with cytokine release. Ageing is perceived as a process of impaired immune functions: IL-1β, IL-6 and tumour necrosis factor-alpha (TNF-α) secretion are increased while IL-2 release is reduced in advanced age. At present, little information is available about perioperative immune reactions at different stages of ageing. The aim of the present study was to compare IL-6, IL-1β, TNF-α, IL-10 and soluble IL-2 receptor (sIL-2R) in younger and older patients undergoing cardiac surgery. Male patients (n = 14) undergoing elective coronary artery bypass grafting (CABG) surgery employing CPB with moderate hypothermia were divided into two groups according to their age: group 1 included seven patients < 50 years old, group 2 included seven patients > 65 years old. All patients received general anaesthesia using a balanced technique with sufentanil, isoflurane and midazolam. Blood samples were collected pre-operatively (T1); intra-operatively during CPB (T2); post-operatively on the day of surgery (T3); on the first post-operative day (T4). Blood concentrations of IL-6, IL-1β, IL-10, TNF-α and sIL-2R were measured using commercially available ELISA kits and corrected for plasma cell volume. Statistical analysis was performed by non-parametric analysis of variance and Mann–Whitney U-test. Significance level was set to P < 0·05. There were no statistically significant differences in the perioperative release of TNF-α, IL-6, IL-1β, IL-10 and sIL-2R among the two groups. We conclude that the perioperative course of cytokine release in patients undergoing CABG surgery with CPB and comparable perioperative management does not significantly differ in the two age groups.
In vitrodifferentiation of peripheral blood T cells towards a type 2 phenotype is impaired in rheumatoid arthritis (RA)
Authors: Asselin S , Conjeaud H , Fradelizi D , Breban M
Published: March 2003

We have examined the capacity of peripheral blood T cells from RA patients to be polarized in vitro towards a type 1 (T1) or a type 2 (T2) phenotype. Peripheral blood T cells from RA patients and from healthy donors were primed by 1 week of culture with soluble OKT3 in the presence of polarizing cytokines. The recovered T cells were restimulated and their cytokine secretion profile determined. Priming of T cells from RA patients in the presence of recombinant (r)IL-2 plus rIL-12 induced a shift towards a T1 pattern, characterized by increased production of interferon-gamma, that was more pronounced than in the case of healthy donors. Conversely, priming of T cells from RA patients in the presence of IL-4 failed to induce a shift towards a T2 profile after 1 week, whereas it induced T cells from healthy donors to acquire such a profile characterized by heightened production of IL-4, IL-5 and IL-13. However, a T2 polarization profile emerged in T cells from RA patients that were primed in the presence of rIL-4 and subsequently maintained in culture in rIL-2 alone for 1 or 2 additional weeks. We conclude that in vitro differentiation of peripheral T cells towards a type 2 phenotype is impaired in RA. Nevertheless, conditions required to drive peripheral T cells towards a type 2 phenotype were established. Administration of autologous polyclonal T cells expressing a type 2 cytokine secretion profile is proposed as a therapeutic strategy in RA.
An anti-CD19 antibody coupled to a tetanus toxin peptide induces efficient Fas ligand (FasL)-mediated cytotoxicity of a transformed human B cell line by specific CD4+ T cells
Authors: Eberl G , Jiang S , YU Z , Schneider P , Corradin G , Mach J-P
Published: March 2003

Treatment of B cell lymphoma patients with MoAbs specific for the common B cell marker (CD20) has shown a good overall response rate, but the number of complete remissions is still very low. The use of MoAbs coupled to radioisotopes can improve the results, but induces undesirable myelodepression. As an alternative, we proposed to combine the specificity of MoAbs with the immunogenicity of T cell epitopes. We have previously shown that an anti-Igλ MoAb coupled to an MHC class II-restricted universal T cell epitope peptide P2 derived from tetanus toxin induces efficient lysis of a human B cell lymphoma by a specific CD4+ T cell line. Here we demonstrate that the antigen presentation properties of the MoAb–peptide conjugate are maintained using a MoAb directed against a common B cell marker, CD19, which is known to be co-internalized with the B cell immunoglobulin receptor. In addition, we provide evidence that B cell lysis is mediated by the Fas apoptosis pathway, since Fas (CD95), but not tumour necrosis factor receptor (TNFr) or TNF-related receptors, is expressed by the target B cells, and FasL, but not perforin, is expressed by the effector T cells. These results show that B cell lymphomas can be ‘foreignized’ by MoAb–peptide P2 conjugates directed against the common B cell marker CD19 and eliminated by peptide P2-specific CD4+ T cells, via the ubiquitous Fas receptor. This approach, which bridges the specificity of passive antibody therapy with an active T cell immune response, may be complementary to and more efficient than the present therapy results with unconjugated chimeric anti-CD20 MoAbs.
Inhibition of IL-12 synthesis of peripheral blood mononuclear cells (PBMC) stimulated with a bacterial superantigen by pooled human immunoglobulin: implications for its effect on Kawasaki disease (KD)
Authors: Takata Y , Seki S , Dobashi H , Takeshita S , Nakatani K , Kamezawa Y , Hiraide H , Sekine I , Yoshioka S
Published: March 2003

The aim of this study was to further assess the role of pooled human immunoglobulin (PHIG) on cytokine production from PBMC stimulated with a bacterial superantigen. Human PBMC were cultured with Streptococcus pyrogenic exotoxin A (SPE-A) with or without PHIG and several proinflammatory cytokine levels of culture supernatants were measured. Serum cytokine levels of KD patients before and after PHIG therapy were also examined. PHIG greatly reduced the production of IL-12, interferon-gamma (IFN-γ) and other cytokines from SPE-A-stimulated PBMC, while exogenous IL-12, but neither IL-1 nor tumour necrosis factor-alpha (TNF-α), restored IFN-γ production inhibited by PHIG. Although PHIG partially adsorbed SPE-A, its inhibitory effect on cytokine production was not played by anti-SPE-A antibody. Although purified CD4+ T cells cultured with human HLA-DR-transfected mouse L cells and SPE-A could not effectively produce IFN-γ, they produced large amounts of IFN-γ if exogenous IL-12 was introduced. KD patients in the acute phase had higher levels of serum IFN-γ than did controls and patients with bacterial infection. Although IL-12 levels of children with or without KD were not significantly different, IL-12 levels of children were much higher than those of adults. However, serum levels of IL-12 of KD patients were transiently but significantly decreased by PHIG therapy and IFN-γ amounts subsequently reverted to basal levels thereafter. These findings indicate that PHIG inhibits IL-12 production of SPE-A-activated monocytes and thereby decreases IFN-γ synthesis by T cells and suggest that inhibition of IL-12 and IFN-γ production is an important part of the mechanisms underlying PHIG therapy on KD.
Adhesion molecules in common variable immunodeficiency (CVID)—a decrease inl-selectin-positive T lymphocytes
Authors: Nordøy I , Müller F , Aukrust P , FRøland S S
Published: March 2003

CVID is immunologically characterized by defective antibody production. Various additional immunological abnormalities have been reported, but little is known of the role of adhesion molecules in CVID. In 31 CVID patients serum levels of l-selectin (CD62L), vascular cell adhesion molecule-1 (VCAM-1) (CD106) and intercellular adhesion molecule-1 (ICAM-1) (CD54) were significantly elevated compared with controls. In 15 CVID patients investigated, the number of l-selectin-positive cells was significantly reduced in both CD4+ and CD8+ lymphocytes compared with controls, and these changes were observed in both CD45RA+  and CD45RO+ subsets. In CD19+ lymphocytes the percentage of ICAM-1+ cells was significantly increased compared with controls. Fifty percent of the patients had splenomegaly. These patients demonstrated even higher serum levels of adhesion molecules, a lower percentage of l-selectin-positive and a higher percentage of CD38+ cells in many T lymphocyte subsets compared with both other CVID patients and controls. Finally, in this patient group the percentage of l-selectin-positive CD19+ lymphocytes was significantly reduced compared with both other patients and controls. These findings indicate a state of ongoing T lymphocyte activation in CVID, especially in the subgroup of patients with splenomegaly, which may contribute to the impaired anti-microbial defence observed in these patients.
A proportion of proteinase 3 (PR3)-specific anti-neutrophil cytoplasmic antibodies (ANCA) only react with PR3 after cleavage of its N-terminal activation dipeptide
Authors: Sun J , Fass D N , Viss M A , Hummel A M , Tang H , Homburger H A , Specks U
Published: March 2003

ANCA directed against PR3 are highly specific for Wegener's granulomatosis and microscopic polyangiitis, and have been implicated in the pathogenesis of small vessel vasculitis. Most PR3-ANCA are directed against conformational epitopes on PR3. This study was designed to determine whether the cleavage of the N-terminal activation dipeptide of PR3 is required for the binding of PR3-ANCA. Recombinant PR3 (rPR3) variants were expressed in the epithelial cell line, 293. As confirmed by radiosequencing, the rPR3 secreted into the 293 cell culture supernatant is N-terminally unprocessed. Two enzymatically inactive rPR3 mutants were expressed in 293 cells: rPR3-S176A and δ-rPR3-S176A. rPR3-S176A contains the N-propetide Ala-2-Glu-1, δ-rPR3-S176A does not. Culture supernatants of rPR3-S176A and δ-rPR3-S176A expressing 293 cells were used as sources of target antigen for PR3-ANCA testing by capture ELISA. Forty unselected consecutive PR3-ANCA+ sera were tested. With δ-rPR3-S176A as antigen all 40 were recognized, compared with only 34 of 40 when rPR3-S176A served as target antigen. The majority of the serum samples contained a mixture of antibodies reacting with epitopes accessible on the mature and on the proform of PR3. In conclusion, the cleavage of the N-terminal activation dipeptide of PR3 is not an absolute requirement for recognition by all PR3-ANCA. However, a substantial proportion of PR3-ANCA recognize (a) target antigen(s) exposed only after the conformational change of PR3 associated with the N-terminal processing. In 15% of sera this PR3-ANCA subset occurred exclusively. PR3-ANCA subtypes can be differentiated using specifically designed rPR3 variants as target antigens, and non-haematopoietic mammalian cells without regulated secretory pathway can be used for their expression.
Cartilage oligomeric matrix protein (COMP)-induced arthritis in rats
Authors: Carlsén S , Hansson A-S , Olsson H , Heinegård D , Holmdahl R
Published: March 2003

In rheumatoid arthritis peripheral cartilaginous joints are inflamed and eroded. One driving factor may be an immune response towards proteins in the cartilage. Here it is shown that cartilage oligomeric matrix protein (COMP), expressed specifically in cartilage, is arthritogenic in the rat. Both native and denatured rat COMP induced severe arthritis in selected rat strains. The arthritis occurred only in peripheral joints which were attacked by an erosive inflammatory process similar to that seen in the human disease. The disease was self-limited and no permanent destruction of joints was seen macroscopically. Disease development appeared to be dependent on an immune response to autologous (rat) COMP and not on cross-reactivity to other cartilage rat collagens (types II, IX and XI). The disease and the immune response to COMP were genetically controlled by the MHC; the RT1u and RT1l haplotypes were more susceptible than the a, c, d, f and n haplotypes. Both LEW and E3 gene backgrounds were highly permissive for disease induction. These findings suggest that the induction of arthritis with rat COMP represents a unique pathogenesis which is controlled by different genes compared with collagen-induced arthritis or adjuvant-induced arthritis.
Imbalances in serum proinflammatory cytokines and their soluble receptors: a putative role in the progression of idiopathic IgA nephropathy (IgAN) and Henoch–Schönlein purpura nephritis, and a potential target of immunoglobulin therapy?
Authors: Rostoker G , Rymer J-C , Bagnard G , Petit-Phar M , Griuncelli M , Pilatte Y
Published: March 2003

Following recent experimental data suggesting an aggravating effect of circulating proinflammatory cytokines on the histological lesions of IgAN, we studied changes in serum proinflammatory cytokines and their soluble receptors and antagonists in patients treated with polyvalent immunoglobulins (15 with severe nephropathy who had indicators of poor prognosis: heavy proteinuria, hypertension, altered renal function and Lee's histological grade III or IV; and 14 with moderate forms of IgAN who had permanent albuminuria > 300 mg/day and < 2000 mg/day, Lee's histological grade II and a glomerular filtration rate > 70 ml/min) in comparison with healthy controls (n = 20) and patients with non-IgA nephritides (n = 50). These were measured by means of specific immunometric assays before and after 9 months of immunoglobulin therapy. Total tumour necrosis factor (TNF) serum and IL-6 levels were elevated in IgAN patients before therapy, relative to controls, and normalized after immunoglobulin therapy. Levels of soluble TNF receptor of type I (sR55) and type II (sR75) increased on immunoglobulin therapy. TNF index α-55,75 used to assess biologically available TNF-α (ratio of total TNF-α divided by levels of soluble TNF receptors sR55 and sR75) was elevated before therapy and was below healthy control values after 9 months of immunoglobulin administration. Levels of serum IL-1 receptor antagonist were low prior to immunoglobulin administration in patients with severe forms of IgAN, and normalized on therapy. Serum interferon-gamma was unmodified. The histological activity index correlated with serum total TNF-α, TNF index α-55,75 and serum IL-6 levels, whereas proteinuria correlated with serum total TNF-α and TNF index α-55,75 but not with serum IL-6. These data suggest that the overproduction of proinflammatory cytokine is unbalanced by their natural antagonists in IgAN and Henoch–Schönlein syndrome. This process may play a role in the progression of the disease and be one of the targets of immunoglobulin therapy.
Circulating soluble factor-inhibiting natural killer (NK) activity of fresh peripheral blood mononuclear cells (PBMC) from inflammatory bowel disease (IBD) patients
Authors: GIACOMELLI R , PASSACANTANDO A , FRIERI G , PARZANESE I , D'ALÒ S , VERNIA P , PIMPO M T , PETRUCCI C , CAPRILLI R , CIFONE M G , TONIETTI G
Published: March 2003

This study was performed in order to assess the cytotoxic activity, both natural (NK) and antibody-dependent (ADCC), of PBMC from 38 IBD patients and correlate it with their clinical features. Cytotoxicity assays were performed using sensitive target cells for NK and ADCC activities. In some experiments, highly purified NK cells, obtained both by Percoll density gradient and by co-culturing non-adherent PBMC with RPMI 8866 feeder cells, were used as effector cells. Furthermore, we evaluated NK cell parameters such as number, surface expression of adhesion molecules (CD11a/CD18, CD49d and CD54) and response to different stimuli. We observed a decreased NK cytotoxicity of PBMC from IBD patients, both in ulcerative colitis (UC) and Crohn's disease (CD), independently of the clinical activity of disease. In contrast, the ADCC lytic activity was within normal range. The lower NK cytotoxic activity observed in our IBD patients cannot be related to a decreased number of NK cells, surface expression of adhesion molecules, defective response to IL-2 and maturative defect. Decreased NK activity was induced in PBMC of controls when serum of patients was added and this was unrelated to monocyte-derived modulating factor(s). Our data show a decreased natural killing by fresh PBMC from IBD patients. This lower activity seems to be unrelated to a primary NK cell defect, since purified NK cells exhibited normal levels of killing. It might be hypothesized that serum factors, possibly derived from lymphocytes, with inhibitory properties on NK activity, might be functionally active in the blood of IBD patients, thus modulating NK activity.
Molecular analysis of rheumatoid factor (RF)-negative B cell hybridomas from rheumatoid synovial tissue: evidence for an antigen-induced stimulation with selection of high mutated IgVH and low mutated IgVL/λ genes
Authors: KRENN V , KÖNIG A , HENSEL F , BEREK C , SOUTO CARNEIRO M M , HAEDICKE W , WANG Y K , VOLLMERS H-P , MÜLLER-HERMELINK H K
Published: March 2003

The mutational pattern of IgVH and IgVL genes from synovial tissue B cell hybridomas (n = 8) of patients (n = 4) with rheumatoid arthritis (RA) was analysed, which had been produced by the electrofusion technique without prior in vitro stimulation. The molecular data were correlated with immunohistopathological data and parameters of local disease activity. The IgVH genes of the B cell hybridomas belonged to the VH3 family (DP42; DP47, n = 2; DP53), the VH1 family (DP75), the VH4 family (DP71) and the VH5 family (DP73); 7/7 IgVH genes showed somatic mutations, the R/S ratio (CDR) was > 3 in 4/7 IgVH genes and the mean R/S ratio of all IgVH genes was 9.3 (CDR) and 1.0 (FR), suggesting an antigen-dependent selection. The IgVL/λ genes belonged to the Vλ1 family (DPL2, DPL5, DPL8nf), the Vλ2 family (DPL11, n = 2) and to the Vλ6 family (IGLV6S1); 6/6 IgVL genes showed somatic mutations, the R/S ratio (CDR) was > 3 in 3/6 IgVL genes and the mean R/S ratio of all IgVL was 3.0 (CDR) and 2.3 (FR), suggesting an antigen-dependent selection. The synovial tissue exhibited germinal centres in the follicles (3/4), with the unique distribution of Ki-M4+ follicular dendritic cells and Ki-67+ proliferating cells and a dominance of IgA+ plasma cells (3/3). All patients were positive for RF in serum and exhibited severe local symptoms (swelling 4/4; warmth 4/4; effusion 2/4), whereas the hybridomas were negative for RF. Since B cell hybridomas showed hypermutation and affinity selection for IgVH and IgVL/λ genes and the patients exhibited severe local symptoms with germinal centres in synovial tissue, this study indicates that an antigen-driven process is behind the B cell expansion in the synovial tissue of clinically affected joints. These mutated B hybridomas were negative for RF, thus suggesting that antigens different from RF are also involved in the local B cell expansion and in the chronic synovitis of RA.
Expression of CD30 mRNA, CD30L mRNA and a variant form of CD30 mRNA in restimulated peripheral blood mononuclear cells (PBMC) of patients with helminthic infections resembling a Th2 disease
Authors: KILWINSKI J , BERGER T , MPALASKAS J , REUTER S , FLICK W , KERN P
Published: March 2003

It has been proposed that CD30, a member of the tumour necrosis factor (TNF) receptor superfamily, is preferentially up-regulated on Th2-type human T cells. In order to investigate a correlation between infection with Echinococcus multilocularis and CD30 expression, we analysed regulation of CD30 mRNA, a variant form of CD30 mRNA (CD30v) and CD30 ligand (CD30L) mRNA expression on PBMC from patients with alveolar echinococcosis (AE) using reverse transcriptase-polymerase chain reaction (RT-PCR). In PBMC of patients with AE as well as healthy donors, spontaneous expression of CD30L mRNA and the CD30v mRNA could be detected. However, the intact form of CD30 mRNA could be detected neither in freshly isolated PBMC of patients nor in PBMC of healthy individuals. Expression of CD30L mRNA and the variant form of CD30 mRNA was frequently detected at individual time points during 72 h of culture of PBMC stimulated with crude Echinococcus antigen. In contrast to CD30v or CD30L mRNA expression, induction of CD30 mRNA expression was detected only in three out of six (50%) healthy donors and in 10 out of 21 (48%) patients with alveolar echinococcosis after 72 h of incubation. As a control, mitogenic stimulation of PBMC of both healthy individuals and infected patients led to expression of intact CD30 mRNA within 24 h of culture. These data demonstrate the different expression of two different forms of CD30 mRNA in PBMC of human individuals. The specific induction of CD30 expression is correlated only in rare cases with the clinical status of patients with AE, indicating the lack of a general induction of CD30 mRNA in this Th2-type-dominated helminthic disease. The data provide further evidence that the CD30 receptor is not an exclusive marker for a Th2-type response.
High IgE in rheumatoid arthritis (RA) patients is complexed with anti-IgE autoantibodies
Authors: MILLAUER N , ZUERCHER A W , MIESCHER S M , GERBER H A , SEITZ M , STADLER B M
Published: March 2003

This study presents data on more than 300 RA and allergic patients analysed for their serum levels of anti-immunoglobulin isotype autoantibodies and IgE. We observed high levels of IgE in sera of RA and allergic patients. Interestingly, we measured significantly higher specific IgE levels against Alternaria but not against nine other allergens in the RA compared with the allergic group. As expected, anti-IgG autoantibodies (rheumatoid factors (RF)) of different isotypes were detected in sera from RA patients only. However, we found increased titres of complexed anti-IgE autoantibodies in all RF+ groups and in the allergic group. These findings may explain why despite elevated IgE levels a decreased prevalence of allergic diseases in RA patients has been observed.
Role of peripheral blood mononuclear cell (PBMC) phenotype changes in the pathogenesis of haemorrhagic fever with renal syndrome (HFRS)
Authors: MARKOTIĆ A , DAŠIĆ G , GAGRO A , SABIONCELLO A , RABATIĆ S , KUZMAN I , ZGORELEC R , SMOLJAN I , BEUS I , ŽUPANC T A , DEKARIS D
Published: March 2003

Hantaviruses cause an important human illness, HFRS. Blood samples from 22 HFRS-positive, six seronegative patients and 15 healthy controls were examined in 1995, during the largest HFRS epidemic in Croatia. Results of double- and triple-colour immunofluorescence analysis showed an increased percentage of cytotoxic T cells (CD3+CD8+) in seropositive patients compared with seronegatives and healthy controls. The majority of seropositive HFRS patients expressed activation and memory antigens on T and B lymphocytes. The percentage of CD23+ and CD21+ B lymphocytes was lower in seropositive patients. HFRS patients had elevated levels of sCD23 and five had elevated total IgE. The increased expression of both early and late T cell activation antigens, e.g. CD25, CD71 and HLA-DR, memory cells and sCD23 positively correlated with biochemical parameters (AST, ALT, urea, α2-globulin) during the acute phase of HFRS. The phenotypic changes observed, especially early and late T cell activation markers, as well as memory cells, could be useful parameters in the evaluation of HFRS course, and prognostic factors of HFRS severity. Additional attention should be paid to liver involvement in the pathogenesis of HFRS.
The pathogenicity of islet-infiltrating lymphocytes in the non-obese diabetic (NOD) mouse
Authors: ABLAMUNITS V , ELIAS D , COHEN I R
Published: March 2003

The aim of the present study was to investigate the pathogenic properties of islet-infiltrating lymphocytes related to the severity of the autoimmune destruction of islet β-cells in the NOD mouse. We analysed the development of insulin-dependent diabetes mellitus (IDDM) produced by adoptive transfer of islet lymphocytes from NOD into NOD.scid mice. Here we show that the transfer was most effective when both CD4+ and CD8+ T cells were present in the infiltrate, but CD4+ T cells alone were sufficient to cause the disease. Islet lymphocytes from both females and males transferred diabetes effectively, but the severity of IDDM was higher when female islet lymphocytes were used. Unexpectedly, the sensitivity of male islets to β-cell damage was greater than that of female islets. Treatment of NOD females with a peptide of heat shock protein (hsp)60, p277, known to protect NOD mice from IDDM, reduced the pathogenicity of the islet lymphocytes. In contrast, administration of cyclophosphamide to males, a treatment that accelerates the disease, rendered the islet lymphocytes more pathogenic. More severe disease in the recipient NOD.scid mice was associated with more interferon-gamma (IFN-γ)-secreting islet T cells of the NOD donor. The disease induced by islet lymphocytes was strongly inhibited by co-transfer of spleen cells from prediabetic mice, emphasizing the regulatory role of peripheral lymphocytes. Thus, the cellular characteristics of the islet infiltrate and the pathogenicity of the cells are subject to complex regulation.
Enhanced expression of CTLA-4 (CD152) on CD4+ T cells in HIV infection
Authors: STEINER K , WAASE I , RAU T , DIETRICH M , FLEISCHER B , BRÖKER B M
Published: March 2003

CTLA-4 (CD152) is a surface molecule of activated T cells with sequence homology to CD28. Both molecules bind to the same ligands, B7.1 (CD80) and B7.2 (CD86) but have antagonistic functions. While CD28 is an important costimulator, CTLA-4 has an essential inhibitory function in maintaining the homeostasis of the immune system. Down- regulation of CD28 predominantly on CD8+ T cells has been described in HIV infection, but analysis of CTLA-4 is complicated by its low expression levels. Here we have used potent signal enhancement to study CTLA-4 on peripheral blood mononuclear cells (PBMC) during HIV infection. CTLA-4 was expressed only on T cells. Expression levels were significantly increased selectively on CD4+ T cells during all stages of HIV infection, while CTLA-4 expression on CD8+ T cells was always low. In contrast, after stimulation with the mitogen phytohaemagglutinin (PHA), CTLA-4 levels were strongly increased on T cells from controls but in T cells from HIV patients this response was severely impaired. Our data suggest that in HIV infection CD4+ and CD8+ T cells may be less responsive to B7 costimuli due to two different mechanisms: increase in CTLA-4 expression by CD4+ cells and down-regulation of CD28 by CD8+ cells.
Role of decidual natural killer (NK) cells in patients with missed abortion: differences between cases with normal and abnormal chromosome
Authors: YAMAMOTO T , TAKAHASHI Y , KASE N , MORI H
Published: March 2003

In order to study the mechanism of abortion, the proportions of NK cells in the peripheral blood and decidual lymphocytes were evaluated in both chromosomally normal and abnormal missed abortions. In normal pregnancy, CD56+16−3−NK cells are a major element of decidual lymphocytes. The percentages of CD56+16−3−NK cells of peripheral lymphocytes in normal pregnancies were not statistically significantly different from those of chromosomally normal and abnormal abortions. In the decidua, the percentages of CD56+16−3− NK cells of decidual lymphocytes showed no statistically significant differences between normal pregnancies and chromosomally abnormal abortions. However, the percentages of CD56+16−3−NK cells of chromosomally normal abortions were lower than those of chromosomally abnormal (P = 0.0025). Moreover, the percentages of CD56+16− NK cells in abortions with normal chromosomes were lower than those in normal pregnancies or abortions with abnormal chromosomes (P = 0.0037, P = 0.0025). However, when the proportion of CD56+NK cells expressing CD16 was evaluated, there were no statistically significant differences in the percentages of CD56+16+ NK cells in normal pregnancies and missed abortions with normal chromosomes and abnormal chromosomes. We conclude that the expression of decidual CD56+16−3− NK cells in missed abortions with normal chromosomes is different from abortions with abnormal chromosomes and that this phenomenon may depend on an abnormal immune response of the maternal side.
Mice with early onset of death (EOD) due to lupus glomerulonephritis
Authors: HONDA S , NEMOTO K , MAE T , KINJOH K , KYOGOKU M , KAWAMURA H , MIYAZAWA S , WEERASHINGHE A , WATANABE H , NARITA J , KOYA T , ARAKAWA M , ABO T
Published: March 2003

Both MRL-lpr/lpr (lpr) and BXSB mice fall victim to autoimmune disease as a function of age. To combine their properties, brother–sister mating of (female lpr × male BXSB)F1 mice was done. Mice for mating were selected according to indicators of early onset of glomerulonephritis and subsequent early death (i.e., EOD). This mating was continued for more than 16 generations. The EOD mice thus established had homozygous H-2k/k, lpr/lpr, and possible yaa/- (in the case of males). The average life span of males was 83 days while that of females was 126 days. After 12 weeks of age, the majority (> 80%) of male EOD mice were characterized by the abnormality of urine due to glomerulonephritis. We then characterized how glomerulonephritis was evoked, especially in terms of expanding lymphocyte subsets in various immune organs. Similar to the case of parental lpr mice, the major expanding cells were CD4−8−B220+ TCRint cells in the immune organs and kidney. In addition, myeloid cells were found to infiltrate the kidney. This massive infiltration of both TCRint cells and myeloid cells might be responsible for the onset of acute glomerulonephritis. Even after more than 50 generations, these EOD mice still carry both lpr and yaa genes. These results suggest that EOD mice might be a very useful tool for the study of acute lupus glomerulonephritis which is evoked by the genetic abnormalities.
Vitamin D, thyroid hormones and muscle mass influence natural killer (NK) innate immunity in healthy nonagenarians and centenarians
Authors: MARIANI E , RAVAGLIA G , FORTI P , MENEGHETTI A , TAROZZI A , MAIOLI F , BOSCHI F , PRATELLI L , PIZZOFERRATO A , PIRAS F , FACCHINI A
Published: March 2003

Increasing evidence has demonstrated that the immune system closely interacts with other physiological systems, whose communications are mediated by circulating cytokines and hormones. The aim of our study was to test whether the number and cytolytic activity of NK cells in a group of relatively healthy Italian nonagenarians and centenarians were affected by the modifications of endocrine, metabolic and functional parameters that occur during ageing. Because of the extreme age of the study population, a cross-sectional analysis was performed. This study revealed that the group of oldest subjects with the highest number of NK cells and the best preserved cytolytic function also presented a preserved metabolism of thyroid hormones and vitamin D and integrity of muscle mass. In fact, the NK cell number and/or cytolytic activity of healthy subjects > 90 years old was positively associated with serum levels of vitamin D, while T3, FT4, i-PTH hormones and lean body mass were associated only with NK cell number. In conclusion, our results stress the paramount importance of nutritional evaluation in the clinical assessment of elderly people.
Lactobacilli from human gastrointestinal mucosa are strong stimulators of IL-12 production
Authors: HESSLE C , HANSON L Å , WOLD A E
Published: March 2003

Interaction of macrophages with bacteria is a stimulus for production of cytokines such as IL-10 and IL-12. IL-12 stimulates T cell and natural killer (NK) cell cytotoxicity and interferon-gamma (IFN-γ) production. IL-10 opposes the T cell-stimulating action of IL-12, decreases the release of proinflammatory cytokines from macrophages, and stimulates B cells. We have studied the capacity of human intestinal isolates from the three Lactobacillus species dominating on the human gastrointestinal mucosa, L. plantarum, L. rhamnosus and L. paracasei ssp. paracasei, to induce production of IL-10 and IL-12 from human blood mononuclear cells, or monocytes. Whole killed lactobacilli were potent stimulators of IL-12 over a wide range of bacterial concentrations. Lactobacillus paracasei gave the highest levels of IL-12 (1.5 ng/ml in response to 5 × 106 bacteria/ml), roughly 10 times more than obtained by stimulation with L. rhamnosus or L. plantarum. Escherichia coli induced on average < 50 pg/ml of IL-12 regardless of the bacterial concentration used. The secretion of free p40 subunit IL-12 followed the same pattern as the secretion of p70 (bioactive IL-12) with regard to the efficiency of different bacteria as stimulators. Escherichia coli was the most efficient trigger of IL-10 production, inducing 0.5 ng/ml IL-10 after stimulation with 5 × 106 bacteria/ml. Lactobacillus rhamnosus induced the highest levels of IL-10 among the lactobacilli (0.5 ng/ml) compared with 0.1 ng/ml evoked by L. plantarum or L. paracasei, but 10 times more bacteria were required for optimal stimulation than with E. coli. When neutralizing anti-IL-10 antibodies were added to the cultures, the IL-12-inducing capacity of L. rhamnosus was increased markedly, while that of E. coli remained low. The results show that mucosa-associated lactobacilli can be potent stimulators of IL-12, and thus potentially of cell-mediated immunity, if they pass over the gut epithelial barrier and interact with cells of the gut immune system.
C3 and C4 allotypes in anti-neutrophil cytoplasmic autoantibody (ANCA)-positive vasculitis
Authors: PERSSON U , TRUEDSSON L , WESTMAN K W A , SEGELMARK M
Published: March 2003

In ANCA-associated small vessel vasculitis few genetic factors have proven to be of importance for disease susceptibility, an exception being deficiency of α1-anti-trypsin, the main inhibitor of proteinase 3 (PR3). Alerted by our finding that myeloperoxidase has affinity for C3, and the finding of an increased frequency of the C3F allele in systemic vasculitis in a British cohort, we examined polymorphism of C3 and C4 in patients with ANCA+ small vessel vasculitis. After identification of all patients at our department with a positive ANCA test during the period 1991–95 and a diagnosis of small vessel vasculitis, blood samples were collected after informed consent. The 67 included patients were grouped according to ANCA serology and disease phenotype using the Chapel Hill nomenclature. The gene frequency of C3F was found to be increased (0.32) compared with controls (0.20; P < 0.05) in the PR3-ANCA+ subgroup. The frequency of C4A3 was increased in the group as a whole, but no increase of C4 null alleles was seen. The findings imply a role for the complement system in the pathogenesis of ANCA-associated small vessel vasculitis.
Use of a whole blood assay to evaluatein vitroT cell responses to new leprosy skin test antigens in leprosy patients and healthy subjects
Authors: WEIR R E , BRENNAN P J , BUTLIN C R , DOCKRELL H M
Published: March 2003

Development of an immunological tool to detect infection with Mycobacterium leprae would greatly benefit leprosy control programmes, as demonstrated by the contribution of the tuberculin test to tuberculosis control. In a new approach to develop a ‘tuberculin-like’ reagent for use in leprosy, two new fractions of M. leprae depleted of cross-reactive and immunomodulatory lipids—MLSA-LAM (cytosol-derived) and MLCwA (cell wall-derived)—have been produced in a form suitable for use as skin test reagents. T cell responses (interferon-gamma (IFN-γ) and lymphoproliferation) to these two new fractions were evaluated in a leprosy-endemic area of Nepal using a simple in vitro whole blood test. The two fractions were shown to be highly potent T cell antigens in subjects exposed to M. leprae—paucibacillary leprosy patients and household contacts. Responses to the fractions decreased towards the lepromatous pole of leprosy. Endemic control subjects also showed high responses to the fractions, indicating high exposure to M. leprae, or cross-reactive mycobacterial antigens, in this Nepali population. The new fractions, depleted of lipids and lipoarabinomannan (LAM) gave enhanced responses compared with a standard M. leprae sonicate. The cell wall fraction appeared a more potent antigen than the cytosol fraction, which may be due to the predominance of the 65-kD GroEL antigen in the cell wall. The whole blood assay proved a robust field tool and a useful way of evaluating such reagents prior to clinical trials.
Expression of mucosa-related integrin αEβ7 on alveolar T cells in interstitial lung diseases
Authors: LOHMEYER J , FRIEDRICH J , GRIMMINGER F , MAUS U , TENTER R , MORR H , VELCOVSKY H G , SEEGER W , ROSSEAU S
Published: March 2003

The expression of αEβ7 integrin has been related to the selective retention of lymphocytes in mucosal tissues of gut, urogenital tract and lung. To identify potential disease-associated αEβ7 expression patterns on cells accounting for lymphocytic alveolitis in interstitial lung disease (ILD),αE expression on CD4+ and CD8+ T cell subsets was evaluated by dual-colour flow cytometry in peripheral blood and bronchoalveolar lavage fluid (BALF) of patients with idiopathic pulmonary fibrosis (IPF; n = 18), hypersensitivity pneumonitis (HP; n = 20) and sarcoidosis (n = 44) in comparison with healthy controls (n = 15). In both healthy individuals and all patient groups the proportion of αE-bearing T cells in peripheral blood was < 2%, whereas the vast majority of alveolar CD8+ T cells consistently co-expressed αE. Absolute alveolar CD8+αE+ cell numbers/ml were up to 30-fold increased in HP patients. Proportions of αE-bearing CD4+ cells in BALF were significantly elevated in IPF (74.0 ± 2.7%) and HP (70.0 ±  2.4%) compared with normals (30.0 ± 1.8%) (mean ±  s.e.m.; P < 0.01). In sarcoidosis, the αE expression on BALF CD4+ cells displayed subgroup dependency: proportions significantly lower than normal were noted in chest radiographic stage I (14.3 ± 1.5%), but increased proportions in stages II (50.0 ± 3.8%) and III (64.0 ± 4.8%). Correlations between common markers of T cell activation or BALF transforming growth factor-beta (TGF-β) bioactivity and αE expression were not noted. We conclude that the vast majority of alveolar CD8+ T cells consistently express αEβ7 and that distinct patterns of αEβ7 expression on alveolar CD4+ lymphocytes in sarcoidosis are related to the diverse manifestations of the sarcoid inflammatory process in the lung.
Leflunomide protects mice from multiple low dose streptozotocin (MLD-SZ)-induced insulitis and diabetes
Authors: STOSIC-GRUJICIC S , DIMITRIJEVIC M , BARTLETT R
Published: March 2003

In certain animal models of autoimmunity the isoxasol derivative leflunomide has been reported to exert a protective effect against autodestruction. In the present study, the immunomodulatory potential of the main metabolite of leflunomide, A77 1726, in experimentally induced autoimmune diabetes was investigated. The disease was induced in genetically susceptible CBA/H mice by multiple low doses of streptozotocin (MLD-SZ, 40 mg/kg per day, given intraperitoneally for 5 consecutive days). Effects of leflunomide were evaluated by two treatment protocols: mice treated with MLD-SZ were injected intraperitoneally with A77 1726 for 10 consecutive days, either during the first 10 days of the disease (early treatment), or starting from day 10 after disease induction (late treatment). Disease manifestations defined by hyperglycaemia, mononuclear infiltration into pancreas, expression of interferon-gamma (IFN-γ) and inducible nitric oxide synthase (iNOS) and destruction of the islets of Langerhans were reduced in a dose-dependent fashion after early treatment with A77 1726 (dose range of 5–35 mg/kg per day). Moreover, late treatment with the high dose of the drug (25 mg/kg per day), started when the autoimmune disease was already apparent, arrested progression of ongoing inflammatory response. Analysis of the effects of A77 1726 on the adhesive interactions of spleen-derived or peripheral blood-derived mononuclear cells from MLD-SZ-treated and normal mice demonstrated that the drug inhibits both ex vivo and in vitro spontaneous mononuclear cell aggregation, thus suggesting that an important component of leflunomide's immunomodulatory action is suppression of adhesive interactions. These results demonstrate both preventive and therapeutic effects of leflunomide in a model of MLD-SZ-induced diabetes and suggest that the drug may be considered a potent therapeutic tool for autoimmune inflammatory disorders, including diabetes.
Antibodies to a non-repeat region ofPlasmodium falciparumantigen Pf155/RESA in individuals from malaria-endemic areas
Authors: SIDDIQUE A B , AHLBORG N , WARSAME M , PERLMANN P , BERZINS K
Published: March 2003

Human antibodies to the repeat regions of the Plasmodium falciparum asexual blood stage antigen Pf155/RESA interfere with parasite growth in vitro, but the significance in this respect of antibodies to non-repetitive epitopes is less clear. In this study the levels of antibodies to a non-repetitive part of Pf155/RESA (residue 199–221) in malaria-exposed individuals were analysed, as was the parasite-inhibitory capacity of such antibodies. Residue 199–221 is of particular interest since it includes a sequence homologous to a cytoadherence-related motif from band 3. Sera from donors in Liberia and Tanzania were analysed for reactivity in ELISA with synthetic peptides together overlapping this part of Pf155/RESA. High antibody reactivity was observed in most of the sera with two peptides including residues 199–211 and 202–214, respectively. Specific antibodies were affinity-purified from selected sera using these peptide sequences and were shown to react with Pf155/RESA by immunofluorescence and Western blotting. The purified antibodies were furthermore shown to inhibit parasite growth in vitro. The results suggest that both repeat and non-repeat epitopes in Pf155/RESA elicit antibodies with potential to protect against malaria infection.
Heat treatment of normal human sera reveals antibodies to bactericidal permeability-inducing protein (BPI)
Authors: BROWNLEE A A , LOCKWOOD C M
Published: March 2003

Heat treatment of normal sera to 56°C for 30 min, a common procedure for the inactivation of viruses, e.g. HIV, reveals the presence of antibodies to neutrophil cytoplasm antigens (ANCA), as detected by indirect immunofluorescence on ethanol-fixed human neutrophils and by antigen-specific ELISA for BPI. Reactivity was not seen to the other common vasculitis-associated antigens proteinase 3 (PR3) or myeloperoxidase (MPO). The effect of temperature was maximal at 56°C, with substantial antibody demonstrable after only 5 min at this temperature. In experiments using polyethylene glycol (PEG)6000 to remove immune complexes, the effect of heating could be abrogated by preincubation with 8% PEG, which suggested that these anti BPI antibodies might be complexed in sera. After passage of normal plasma over a protein G column, the acid-eluted fraction contained elevated levels of antibodies to BPI but not to other vasculitis-associated antigens such as PR3 or MPO, nor to glomerular basement membrane (GBM), the Goodpasture antigen which is recognized by the pathogenically important human antibodies shown to mediate nephritis in transfer experiments. Moreover the levels of anti-BPI in the IgG fraction could be augmented by preincubation with glycine pH 2.5 for 30 min. This anti-BPI activity could be inhibited by addition of the unbound material from the protein G column and this inhibitory material was not heat-labile at 56°C. The molecular specificity of this autoreactivity was confirmed using recombinant BPI in coincubation experiments and the epitope localized to the C or N terminal moieties by the use of recombinant fusion proteins.
Spleen autotransplantation provides restoration of functional splenic lymphoid compartments and improves the humoral immune response to pneumococcal polysaccharide vaccine
Authors: Leemans R , Harms G , Rijkers G T , Timens W
Published: March 2003

After splenectomy, patients have an increased risk of overwhelming post-splenectomy infection (OPSI) or sepsis involving encapsulated bacteria such as pneumococs. The value of spleen autotransplantation after splenectomy because of trauma has long been questioned. Much attention has been given to the restoration of mononuclear phagocyte system (MPS) function, which appeared to be similar to that of splenectomized individuals. The presence of specific anti-pneumococcal antibodies may enhance phagocytosis of opsonized bacteria by other parts of the MPS, as present in the liver. Therefore, in the present study we have evaluated the restoration of the humoral immune response after spleen autotransplantation, especially to pneumococcal capsular polysaccharides (PPS). Wistar rats were divided into three groups which were operated as follows: splenectomy, splenectomy followed by autotransplantation, and sham operation. After 12 weeks the rats were vaccinated with 23-valent pneumococcal vaccine. Blood samples were taken after 3 days, 3 and 6 weeks for anti-PPS IgM and IgG ELISA against types 3, 4, 6, 9, 14 and 23. In addition, immunohistological studies were performed on the autotransplants. Significant antibody titre rises were found in a main proportion of the autotransplanted rats, comparable to sham-operated rats. Splenectomized rats showed as well a significantly lower increase in immunoglobulin levels, as significant differences in the proportion of rats showing a minimum two-fold increase of antibody level, considered to represent an adequate response. The titres were highest 3 days after vaccination. Immunohistochemical studies demonstrated structurally functional autotransplants, including an intact marginal zone. Considering this significant anti- pneumococcal antibody response, spleen autotransplants can be expected to enable an improved humoral response to PPS, and to contribute to protection against OPSI after splenectomy.