bluuebunny/crossref_metadata_2025 huggingface dataset:
Cyclin B1 expression in response to abrogation of the radiation‐induced G2/M block in HeLa cells
Authors: Theron T. , Böhm L.
Published: December 2004

The G2 block is a major response of cells to DNA damage and seem to be induced independently of p53 status. It is thought that the G2 block has a protective function and allows cells to repair their DNA. The molecular events involved in the formation of the G2 block therefore are of great interest. We have used pentoxifylline, a potent G2 delay abrogator, to study the expression of an essential component of the mitosis promoting complex (MPF), cyclin B1. Cyclin B1/G2 ratios are used to show that irradiation induces a decrease in cyclin B1 expression and that pentoxifylline restores cyclin B1 expression to control level. This confirms that suppression of cyclin B1 plays a role in the formation of the G2 cell cycle delay, and that elevating cyclin B1 expression is part of the mechanism of action of pentoxifylline on G2 blocked cells.
Sulindac enhances cell proliferation in DMH‐treated mouse colonic mucosa
Authors: Moorghen M. , Orde M. , Finney K. J. , Appleton D. R. , Watson A. J.
Published: December 2004

In a previous study we reported that the NSAID sulindac had a marked inhibitory effect on the development of colonic tumours in mice treated with the carcinogen 1,2‐dimethylhydrazine (DMH). In this study we examined the effects of sulindac in respect of cell‐kinetic changes in mouse colonic mucosa as determined by flash labelling with the thymidine analogue bromodeoxyuridine (BrdUrd) at varying intervals during the process of colonic carcinogenesis. We also investigated the possibility that these changes may be modulated by misoprostol a prostaglandin E1 analogue. Four groups of 36 mice each were treated for 18 weeks with the following drug/s respectively: (1) DMH; (2) DMH and sulindac; (3) DMH, sulindac and misoprostol; and (4) DMH and misoprostol. Three animals from each group were killed each week between the sixth week and the eighteenth week after the start of the experiment. A 1‐h flash label technique was employed and paraffin sections of colonic mucosa were examined. For each animal a total of 50 perfect axially cut crypts were chosen and the following parameters determined: crypt length, labelling index and labelling index distribution: the data were analysed using the computer program GLIM. For each of the four groups, crypt lengths increased significantly with the duration of treatment with no significant difference between the groups. In sulindac‐treated animals the labelling index for all positions increased with duration of treatment whereas for animals not treated with sulindac there was no significant difference in labelling index with respect to duration of treatment. The administration of misoprostol did not appear to significantly alter the effects of sulindac. It is postulated that the observed increase in cell proliferation could be a compensatory phenomenon occurring secondary to loss of crypt epithelial cells by apoptosis induced by sulindac. Also the finding of an increase in labelling index mediated by a chemopreventive agent indirectly questions the rationale behind the therapeutic manipulation of crypt cell proliferation in order to reduce the risk of colon cancer.
Promotion‐sensitive epidermal and mammary epithelial cells maintained in suspension over agarose
Authors: Eto I.
Published: December 2004

The molecular basis of tumour promotion is still largely unknown. In in vitro model of tumour promotion, the promotion‐sensitive cells are induced to grow under anchorage‐independent conditions in the presence of promoting agent. The customary way of providing such conditions is to immobilize these cells in soft agar, but such cells cannot be readily recovered to study the induced biochemical and molecular events. In the present report, we analysed these events using JB6 mouse epidermal cells maintained in suspension in liquid medium over agarose. 12‐O‐Tetradecanoylphorbol‐13‐acetate (TPA) induced anchorage‐independent synthesis of DNA in promotion‐sensitive P+ (but not in promotion‐resistant P−) JB6 cells and this TPA‐induced synthesis of DNA positively correlated with TPA‐induced formation of colonies in soft agar. The TPA‐induced synthesis of DNA began on or shortly before 24 h after the introduction of TPA, peaked at about 48 h and then declined to the control levels over the next several days. All trans‐retinoic acid and dexamethasone inhibited and calcitriol (1α,25‐dihydroxyvitamin D3) synergistically stimulated this TPA‐induced DNA synthesis. Western immunoblot analysis of cyclins (A, B1, D1 and E) and p27Kip1, a cyclin‐dependent kinase inhibitor, indicated that TPA induced cyclin A and cyclin B1 expression in P+ (but not in P−) JB6 cells and this induction coincided in time with TPA‐induced synthesis of DNA. TPA also strongly induced cyclin D1 expression in P+ (but not in P−) JB6 cells, but this induction started prior to the expression of cyclin A and cyclin B1. TPA did not affect the expression of either cyclin E or p27Kip1 to any significant extent. We also found that NMU38 rat mammary epithelial cells were operationally equivalent to the promotion‐sensitive P+ JB6 cells, but in these cells 17β‐oestradiol exerted a strong synergistic effect on TPA‐induced synthesis of DNA. Based on these observations, we tentatively propose a sequence of molecular events which possibly lead to the anchorage‐independent synthesis of DNA in these cells.
Who Takes Care of Education? Looked after children's perceptions of support for educational progress
Authors: Harker Rachael M , Dobel‐Ober David , Lawrence Julie , Berridge David , Sinclair Ruth
Published: December 2004

This paper documents the views of a sample of 80 children and young people, aged 10–18 years, living in foster and residential care placements in England, regarding their educational experience whilst being looked after. The children were interviewed as part of an ongoing evaluation of the Taking Care of Education project, a development programme designed to facilitate improvements in the education of looked after children, co‐ordinated by the National Children's Bureau and funded by the Gatsby Charitable Foundation. Interviews with the sample asked children to assess their current educational progress and identify individuals who supported or hindered their education, as well as the availability of educational support in care placements. Teaching staff were frequently mentioned as providers of support, whilst social workers were often associated with hindering educational progress. Children and young people had access to a range of educational supports in care placements, and these were more widely available in residential settings than in foster care. Children offered a range of pertinent suggestions as to how the educational experience of those in the public care system might be improved.
Protecting children in Scotland: examining the impact of the Children (Scotland) Act 1995
Authors: McGhee Janice , Francis Joe
Published: December 2004

This paper discusses findings from a small‐scale study of the impact on child protection practice following implementation of the Children (Scotland) Act 1995. The Act introduced three new measures to allow the state to intervene in families to protect a child where there is a risk of significant harm. These include the child protection order, the child assessment order and the exclusion order. The child protection order provides for the removal of a child to (or his or her retention in) a place of safety. In the first two years of the operation of the 1995 Act fewer applications were made for this order compared with similar provisions under the previous legislation. This reduction in applications appears to be related to unfamiliarity with new legislation; greater scrutiny resulting from the more formal application to the sheriff court; and the introduction of a new legal criterion for intervention, the presence or likelihood of ‘significant harm’. The introduction of the ‘no order’ principle into Scottish child care law is also likely to be a factor.
Informing parents of visually impaired children: who should do it and when?
Authors: Speedwell L. , Stanton F. , Nischal K. K.
Published: December 2004

Background  Parents of sick or disabled children are likely to be more stressed than parents of non‐disabled children and may benefit from being given information about their child's condition and its implications, but the stage at which parents should receive such information and who should provide it has not been fully investigated.

The impact of written information on stress levels of parents of visually impaired children, seen in the ophthalmology clinic of a children's hospital, was explored in this study and the question of who parents thought should provide information, and at what stage, was also investigated.

Methods  A longitudinal, experimental intervention study was conducted to compare the effect on perceived stress levels of providing information about the implications of visual impairment to parents. Effects were compared according to the child's age and explored in relation to baseline stress levels. The study included a frequency analysis about parents’ knowledge of visual impairment and how it relates to education, and their response to being given written information.

Results  The results did not show an effect on levels of parental stress but did find that parents of school age children were more stressed than those of preschool age. Over 80% of participants considered that information was given too late and suggested it should be given soon after diagnosis. Of the controls, 32.6% thought the general practitioner should provide information on education although participants were more likely to expect the hospital to provide it.

Conclusions  The majority of parents would prefer to receive information soon after diagnosis of their child's visual impairment. Most parents do not know who to approach for information on education. Giving them access to patient liaison teams who could advise about the repercussions of visual impairment in children would be beneficial.
Which factors explain the lower prevalence of atopy amongst farmers' children?
Authors: Remes S. T. , Iivanainen K. , Koskela H. , Pekkanen J.
Published: December 2004

Background The inverse association between farming and atopy in children has been attributed to microbial exposure, especially through livestock. Very little is known about other potential explanatory factors.

Objective To explore potential differences in lifestyle and environmental factors between farmer and non‐farmer families, and whether these factors could explain the association between farming and childhood atopy.

Methods A cross‐sectional study, including 366 farmers' and 344 non‐farmers' children in eastern Finland. Information regarding exposure and background characteristics was gathered by a written questionnaire. Atopy was defined as having one or more positive skin prick test reactions (> 3 mm) against the six common aeroallergens.

Results Regardless of the current farming type, atopy was less frequent among the farmers' children than the non‐farmers' children (aOR 0.56, 95% CI 0.40–0.78). Remarkable differences were seen in many lifestyle factors (including diet) between the farmer and non‐farmer families, but only a few of the explored factors were associated with atopy. The frequency of current livestock contacts seemed to have an inverse, dose–dependent association with atopy (aOR 0.46, 95% CI 0.22–0.97 for daily vs. no contact). Having lived on a dairy farm in infancy (aOR 0.51, 95% CI 0.28–0.93), or having had cats or dogs in infancy (aOR 0.60, 95% CI 0.42–0.85), decreased the risk of atopy at school age. The inverse association between farming and atopy was not explained by the sociodemographic factors, or by differences in conventional risk factors of atopy. Animal contacts explained partially, but not completely, the association.

Conclusion Higher frequency of animal contacts is one factor, but probably not the only one, explaining the inverse association of farming and atopy in children. The importance of early life exposures may have recently been over‐emphasized, and current exposures discounted, when studying the risk factors of childhood atopy.
Developmental changes in interleukin‐12‐producing ability by monocytes and their relevance to allergic diseases
Authors: Itazawa T. , Adachi Y. , Okabe Y. , Hamamichi M. , Adachi Y. S. , Toyoda M. , Morohashi M. , Miyawaki T.
Published: December 2004

Background The T helper type‐2 (Th2)‐dominated situation can be observed in allergic diseases such as asthma or atopic dermatitis. A reduced ability to produce IL‐12, which is a key cytokine for the induction of Th1 responses, has been proposed to lead to aberrant Th2 development in these disease conditions.

Objective This study was intended to examine how IL‐12‐producing ability might associate with allergic diseases as a function of age.

Methods IL‐12 production by monocytes at various ages was assessed in patients with bronchial asthma and/or atopic dermatitis (n = 100) in comparison with non‐allergic control subjects (n = 144). Whole blood cells were stimulated with lipopolysaccharide (LPS) after priming with IFN‐γ, then intracellular cytokine expression of IL‐12 and IL‐8 as a control cytokine of CD14‐positive cells was assessed by flow cytometric analysis.

Results In the control subjects, the ability of monocytes to produce IL‐12 was negligible at birth and gradually increased with advancing age, whereas IL‐8 production was intense throughout the human life. At more than 7 years of age, IL‐12 production of patients with allergic diseases was significantly lower compared with that of control subjects. The unexpected finding was that infants and children below 6 years of age with allergic diseases tended to produce more IL‐12 compared with age‐matched controls. In this young group, it was noted that enhanced IL‐12 production by monocytes was especially observed in allergic patients with specific IgE antibodies against some food allergens. Significant inverse relationships between serum IgE levels and IL‐12‐producing ability were found in the teenage and adult groups, but not in the younger children.

Conclusion IL‐12 appeared to play different roles in the pathogenesis of allergic diseases between younger and older ages.
Peptide mapping of immunoglobulin E and immunoglobulin G immunodominant epitopes of an allergenic Blomia tropicalis paramyosin, Blo t 11
Authors: Ramos J. D. A. , Cheong N. , Lee B. W. , Chua K. Y.
Published: December 2004

Background The identification of immunodominant peptides containing the IgE and IgG epitopes on allergen molecules is an important step in understanding the interaction of the allergen with the immune system and, thus, essential for the development of effective immunotherapeutic and diagnostic reagents. The present study aimed to map the IgE and IgG immunodominant peptides of Blomia tropicalis (Bt) allergen Blo t 11, a high molecular weight allergen homologous to paramyosin, exhibiting important allergenic activity.

Methods Eleven overlapping fragments of Blo t 11 cDNA gene were expressed as glutathione s‐transferase (GST) fusion peptides, which were affinity‐purified using the glutathione‐Sepharose column. Human IgE and IgG immunodominant peptides were determined by dot blot immunoassay using crude Bt extract‐positive sera from asthmatic patients. Evaluation of allergenicity, specific hIgG subclass analysis, and cross‐ and self‐inhibition studies were determined by enzyme‐linked immunosorbent assay.

Results Blo t 11 contains multiple IgE and IgG immunodominant peptides scattered throughout the molecule. The dominant IgE and IgG peptides were mapped at amino acid positions 336–557 and 698–875, respectively. An immunodominant peptide (fD) registered a higher percentage of IgE and IgG reactivity compared to the rFL‐Blo t 11. Significant serum levels of Blo t 11‐ and fD‐specific IgG1, IgG2 and IgG4, but not IgG3 were detected in the Bt extract‐positive sera tested. Cross‐inhibition study revealed the rFL‐Blo t 11 was significantly inhibited by fD.

Conclusion The IgE and IgG immunodominant peptides of Blo t 11 have been mapped. Our data suggest that utilization of Blo t 11 fragment(s) or chimeric fusion fragments containing IgE and IgG epitopes could be a better alternative in the development of diagnostic and therapeutic reagents for mite allergy.
Par j 1 and Par j 2, the major allergens from Parietaria judaica pollen, have similar immunoglobulin E epitopes
Authors: Asturias J. A. , Gómez‐Bayón N. , Eseverri J. L. , Martínez A.
Published: December 2004

Background Parietaria judaica is the main cause of allergy in Mediterranean countries. The major allergens from P. judaica pollen, Par j 1 and Par j 2, have amino acid sequence identity of 45% and contain eight cysteine residues involved in disulphide bonds that compact the structure.

Objective The aim of the study was to identify IgE‐binding epitopes on Par j 1 and Par j 2, the major allergens from P. judaica pollen and correlate them with the three‐dimensional structure of the proteins.

Methods Overlapping peptides representing the complete length of Par j 1 and Par j 2 were synthesized on a cellulose‐derivatized membrane. Sera from 17 P. judaica‐allergenic patients were used to identify IgE‐binding epitopes. Homology modelling of the three dimensional structure of both allergens was generated using the Swiss‐Model software on the basis of previously reported crystal structures.

Results Five and eight IgE‐binding epitopes were identified on Par j 1 and Par j 2 allergens, respectively. Both proteins belonged to the non‐specific lipid transfer proteins (ns‐LTP) family and therefore a three‐dimensional model of both allergens was constructed on the basis of the maize ns‐LTP crystal structure. Par j 1 and Par j 2 allergens have three similar allergenic epitopes with high homology and identical conformation.

Conclusion Three similar IgE‐epitopes of major allergens from P. judaica have been described. They could be good candidates for designing of IgE haptens as therapeutic agents with reduced anaphylactic side‐effects or for developing hypoallergenic variants of these major allergens.
Prevalence of occupational allergy due to live fish bait
Authors: Siracusa A. , Marcucci F. , Spinozzi F. , Marabini A. , Pettinari L. , Pace M. L. , Tacconi C.
Published: December 2004

Background Larvae of insects and worms, used as live fish bait (LFB), are a common source of allergy in anglers and occupationally exposed workers, but the prevalence and predictors have not yet been investigated.

Objective This study assessed the prevalence and associated factors of occupational allergy in workers exposed to LFB.

Methods We assessed the prevalence of sensitization to LFB and work‐related symptoms (WRS) in 76 workers occupationally exposed to it. All workers completed a case history questionnaire and underwent skin prick tests (SPT) for common aeroallergens and bluebottle (Calliphora vomitoria), bee moth (Galleria mellonella), mealworm (Tenebrio molitor), and gusano rojo (Cilecomadia moorei). Specific IgE were tested in 64 subjects. Two‐thirds of the workers had a high level of LFB exposure.

Results Sensitization to LFB was found in 24 workers (31.6%). Seven subjects (9.2%) reported WRS (asthma in 3, rhinoconjunctivitis in 5, and contact urticaria in 1), and 5 were positive for SPT and serum IgE to one or more LFB extracts. One was also exposed to LFB while fishing. Sensitization to LFB extracts and WRS were strongly associated (Odds Ratio = 6.6, P < 0.05). The 7 subjects with WRS had been exposed longer than asymptomatic subjects with positive skin tests to LFB (P < 0.05). No differences in sex, age, smoking habit, duration or level of exposure, and atopy were detected in symptomatic or in sensitized subjects, compared with normal workers.

Conclusion Sensitization to LFB and WRS are relatively frequent in occupationally exposed workers. No associated factors of occupational allergy to LFB could be detected.
Restricted genetic defects underlie human complement C6 deficiency
Authors: DRAGON-DUREY M A , FREMEAUX-BACCHI V , BLOUIN J , BARRAUD D , FRIDMAN W H , KAZATCHKINE M D
Published: December 2004

Complement C6 homozygous deficiency (C6D) has been rarely observed in Caucasians but was reported at higher prevalence among African-Americans. We report on the molecular basis of C6D in seven unrelated black individuals of North or Central Africa descent who live in France. These patients have presented Neisseria meningitidis infection (four cases), focal and segmental glomerulosclerosis with hyalinosis (one case), systemic lupus erythematosus (one case) or Still's disease (one case). All patients exhibited undetectable antigenic C6 by using a sensitive ELISA assay. An additional four cases of complete C6 deficiency with no associated disease have been characterized after family studies. Exons 6, 7 and 12 have been described recently as the location of molecular defects on the C6 gene in randomly chosen black Americans. Genomic DNA from the seven patients were subjected to direct polymerase chain reaction amplification of these three exons. Nucleotide sequencing analysis of the amplified DNA fragments revealed a homozygous single-base deletion (1936delG) in exon 12 in three cases and four compound heterozygous deletions for a single base in exon 7 (1195delC) or in exon 6 (878delA) associated with the same deletion in exon 12 (1936delG). Our observations further establish the restricted pattern of genetic defects associated with homozygous C6 complement deficiency in individuals of African descent.
Preserved immune system in long-term asymptomatic vertically HIV-1 infected children
Authors: RESINO S , CORREA R , BELLÓN J M , MUÑOZ-FERNÁNDEZ M Á
Published: December 2004

The objective of this study was to study immune system status in long-term asymptomatic (LTA) HIV-1-infected children. A cross-sectional study was used, involving HIV-1-infected children over 7 years of age who were rated into two groups according to their clinical and immunological classification: (a) LTA: 7 asymptomatic HIV-1-infected children in A1; (b) Rapid progressor (RP): 14 age-matched C3 HIV-1-infected children. The control group consisted of 17 age-matched uninfected children. The characterization of CD4+ T-cell subsets was determined by three-colour flow cytometry. The proliferative response and cytokine production by activated peripheral blood T-cells were also measured. IL-7 levels were measured in serum. Thymic production of T-cells was quantified by TCR rearrangement excision circles (TRECs). The LTA children showed similar proliferative responses to PHA, PWM and anti-CD3+ anti-CD28, but lower responses to tetanus toxoid and streptokinase, in comparison with the controls but always higher responses in comparison with the RP group. The production of TNF-α and IFN-γ was similar in the LTA and control groups, and both were higher than the levels in the RP group. The LTA group showed a lower percentage of memory CD4+ T-cells (CD4+ CD45RO+, CD4+ CD45RA-CD62L+) than the control and RP groups. The LTA group also showed lower percentages of CD4+ CD7- cells than the controls. As for naïve CD4+ T-cells (CD4+ CD45RA+ CD62L+), CD4+ CD45RA+ and CD4+ CD62L+ cells, the LTA group showed higher values than the control and RP groups. The LTA group showed higher percentages of CD4+ HLA-DR+ CD38+ than the controls, but lower values than the RP group. In contrast, the LTA group had percentages of CD4+ HLA-DR-CD38+ T-cells higher than both the control and RP groups, whereas CD4+ CD38+ levels were only higher in the LTA group in comparison with the controls. CD4+ HLA-DR+ CD38- and CD4+ HLA-DR+ cell numbers were lower in the LTA group in comparison with the RP group. We found almost normal values of TRECs and IL-7 in the LTA group, but lower values in the RP group. Moreover, we found an inverse relation between TREC levels and IL-7 in plasma from HIV-infected children. Asymptomatic HIV-1 infected children have a well preserved immune system similar to that of control uninfected children in spite of HIV-infection for more than 7 years. Moreover, our results identified new markers of HIV disease, such as TRECs and IL-7, that could be used to monitor disease.
Virus load correlates inversely with the expression of cytotoxic T lymphocyte activation markers in HIV-1-infected/AIDS patients showing MHC-unrestricted CTL-mediated lysis
Authors: SINDHU S T A K , AHMAD R , BLAGDON M , AHMAD A , TOMA E , MORISSET R , MENEZES J
Published: December 2004

Cytotoxic T lymphocytes (CTL) are key players to suppress viral load (VL) but CTL responses become compromised with progression of HIV-infection/AIDS. Some progressors develop MHC-unrestricted CTL with anti-CD4+ cytocidal activity. Immune activation status of these CTL and its significance in disease progression are unknown. To determine the relationship between VL and T cell activation, a cross-sectional study was carried out using blood samples from 13 HIV-1-infected/AIDS patients at various stages of progression and seven age-matched seronegative controls. We examined expression of HLA-DR and CD38 activation markers on purified CTL. MHC-unrestricted killing by these CTL was also evaluated against uninfected, allogeneic CD4+ T cells as well as several human cell lines. The expression of activation markers correlated inversely (rs = − 0·91, P < 0·0001) with VL of the subjects. CTL effectors of these patients killed targets expressing or lacking CD4+, independently of MHC class I recognition. Interestingly, the patients with higher VL showed an increased number of γδTCR-bearing CTL in blood and their MHC-unrestricted killing activity was blocked significantly (P < 0·01) by γδTCR-specific monoclonal antibody. CD3+ T counts of these patients were also consistently subnormal. Inverse correlation between VL and CD8+ T cell activation markers seems to be an indicator of CTL-associated immunopathogenesis in HIV patients with elevated γδCTL in the peripheral blood.
A set of recombinant antigens from Echinococcus granulosus with potential for use in the immunodiagnosis of human cystic hydatid disease
Authors: VIRGINIO V G , HERNÁNDEZ A , ROTT M B , MONTEIRO K M , ZANDONAI A F , NIETO A , ZAHA A , FERREIRA H B
Published: December 2004

Several recombinant clones expressing antigens from Echinococcus granulosus were isolated previously from a parasite cDNA library using cystic hydatid disease (CHD) patients’ sera or rabbit hyperimmune antiserum against a lipoproteic fraction from bovine cyst fluid. Six of these antigens were expressed in Escherichia coli and the purified recombinant proteins were tested in enzyme-linked immunosorbent assay (ELISA) for specific IgG with a panel of sera from patients with surgically confirmed (n = 58) or immunologically diagnosed (n = 71) CHD. Sera from clinically normal individuals (n = 203) and sera from individuals with other helminthic infections (n = 65) were assayed for the assessment of specificity. A cut-off value was determined by receiver-operating-characteristic plots for each antigen. A recombinant antigen B subunit (AgB8/2) presented the highest sensitivity (93·1%), considering the group of sera from patients with CHD surgically confirmed, and specificity (99·5%) and is proposed as the basis for an immunodiagnostic test. The other recombinant antigens tested presented sensitivities between 58·6% and 89·7%, and three of them were considered of complementary value. In subclass-specific ELISA, different IgG isotypes showed dominance in the response for each of the recombinant antigens. There was a clear predominance of IgG4 response for all antigens tested, indicating that this would be the subclass of choice to be assessed for these recombinant proteins.
Quantification of indoleamine 2,3-dioxygenase gene induction in atopic and non-atopic monocytes after ligation of the high-affinity receptor for IgE, FcɛRI and interferon-γstimulation
Authors: VON BUBNOFF D , BEZOLD G , MATZ H , HANAU D , De La SALLE H , BIEBER T
Published: December 2004

Antigen-presenting cells (APCs) are crucial in regulating the outcome of T cell responses. Certain APCs are able to down-regulate T cell proliferation in vitro by inducing the enzyme indoleamine 2,3-dioxygenase (IDO) upon interferon-γ (IFN-γ) stimulation. IDO is the rate-limiting enzyme in the catabolism of the essential amino acid tryptophan. A lack of extracellular tryptophan creates environments in which cells become starved for this amino acid. The high-affinity receptor for IgE, FcɛRI, is the principal receptor for the binding of specific IgE in type I-mediated allergies. We demonstrated recently that IDO is overexpressed in FcɛRI-stimulated monocytes. In the present study, we performed quantification of IDO gene induction after treatment of atopic (FcɛRIhigh) and non-atopic (FcɛRIlow/–) monocytes with IgE/anti-IgE and IFN-γ. By quantitative PCR ELISA, we found IDO molecule induction in atopic monocytes was enhanced about 50-fold over non-atopic monocytes after ligation of FcɛRI. Stimulation with IFN-γ at a concentration of 100 U/ml in culture medium caused an increase in IDO gene copy numbers in atopics of about fourfold over that of non-atopics. This comparative quantification study demonstrates clearly the regulation of IDO gene expression by FcɛRI and discloses differences thereof in atopic and non-atopic cells upon inflammatory stimuli.
Expression of CD80 on Kupffer cells is enhanced in cadaveric liver transplants
Authors: KWEKKEBOOM J , KUIJPERS M A , BRUYNEEL B , MANCHAM S , DE BAAR-HEESAKKERS E , IJZERMANS J N M , BOUMA G J , ZONDERVAN P E , TILANUS H W , METSELAAR H J
Published: December 2004

In experimental animals inhibition of T cell co-stimulation immediately after organ transplantation effectively prevents rejection. We investigated whether the expression of co-stimulatory molecules is enhanced in cadaveric liver transplants, whether their expression is influenced by the transplantation procedure, and whether variation in expression between liver transplants is related to the occurrence of acute rejection. Expression of CD80, CD86 and the macrophage marker CD68 were determined by immunohistochemistry in biopsies from 40 clinical liver transplants obtained at different time-points during the transplantation procedure, and in normal liver tissue obtained from 10 human livers. Expression of CD80 and CD86 on Kupffer cells was graded by comparison with CD68-staining. In a subgroup CD80 and CD86 mRNA was quantified by real-time detection polymerase chain reaction. CD86 was expressed in all liver transplants and normal livers on the majority of Kupffer cells. CD80 was absent or sporadically expressed in normal liver tissue, but in 18 of 40 liver transplants at least one-quarter of Kupffer cells expressed CD80. CD80- and CD86-mRNA and protein expression in liver transplants did not change during the warm ischaemic and reperfusion phases of the transplantation procedure. CD80-expression on Kupffer cells varied strongly between individual donor livers; this variation was, however, not significantly related to the occurrence of acute rejection after transplantation. In conclusion, in nearly half of cold-preserved cadaveric liver transplants an increased proportion of Kupffer cells express CD80 at the time of transplantation in comparison with normal liver tissue. The expression was not further induced by warm ischaemia and reperfusion. However, the observed variation in CD80-expression between liver transplants is not a accurate predictive measure for acute rejection.
Specific stimulation of peripheral blood mononuclear cells from patients with acute myocarditis by peptide-bound flavin adenine dinucleotide (FAD), a naturally occurring autologous hapten
Authors: CICEK G , SCHILTZ E , STAIGER J , NEUMANN F-J , MELCHERS I , BRANDSCH R
Published: December 2004

The tryptic FAD-peptide carrying the flavin in 8α-(N3)histidyl linkage as natural hapten was isolated by HPLC from the bacterial enzyme 6-hydroxy-d-nicotine oxidase. The same flavin protein linkage is found in the mitochondrial succinate dehydrogenase flavoprotein subunit, the predominant flavoprotein with covalently bound FAD in mitochondria of cardiomyocytes. Peripheral blood mononuclear cells (PBMC) were isolated from four patients with acute myocarditis, seven patients with dilated cardiomyopathy (DCM) and from four healthy control individuals. The response of PBMC to the FAD-peptide was evaluated by measuring proliferation ([3H]-dThd incorporation) and cytokine secretion [interferon (IFN)-γ]. PBMC from all patients with acute myocarditis showed positive responses to the FAD-peptide, in contrast to PBMC from patients with DCM or control individuals. Following the recovery of the patients from the acute inflammation of the heart, PBMC no longer exhibited a proliferation response to the FAD-peptide. A chemically synthesized FAD-free peptide with identical amino acid sequence induced no response of PBMC. The results are consistent with a recall response by activated T cells, specific for the normally cryptic mitochondrial flavin-hapten, which may be liberated following cardiomyocyte destruction during the inflammation of the heart.
Myeloid and lymphoid dendritic cells in normal pregnancy and pre-eclampsia
Authors: DARMOCHWAL-KOLARZ D , ROLINSKI J , TABARKIEWICZ J , LESZCZYNSKA-GORZELAK B , BUCZKOWSKI J , WOJAS K , OLESZCZUK J
Published: December 2004

The aim of our study was to estimate the populations of peripheral blood myeloid and lymphoid dendritic cells (CD1c+, BDCA-2+) and the CD1c+ : BDCA-2+ ratio in normal pregnant women and in patients with pre-eclampsia. Fifteen women in the first, second and third trimesters of normal pregnancy, and 25 patients with pre-eclampsia were included in the study. The dendritic cells were isolated from peripheral blood, stained with monoclonal antibodies against blood dendritic cell antigens (anti-CD1c, anti-BDCA-2) and estimated using the flow cytometric method. CD1c+ and BDCA-2+ dendritic cells were present in women during all trimesters of physiological pregnancy and in pre-eclamptic patients. It was observed that the numbers of dendritic cells were significantly lower in the second trimester when compared with the first and third trimesters of normal pregnancy. Furthermore, in the second trimester, CD1c+ : BDCA-2+ ratio was higher than in the other trimesters of physiological pregnancy. All populations of dendritic cells and CD1c+ : BDCA-2+ ratio did not differ in the first and third trimesters of normal pregnancy. The percentage of BDCA-2+ dendritic cells was significantly lower in pre-eclampsia in comparison with healthy women in the third trimester of physiological pregnancy, while CD1c+ : BDCA-2+ ratio was significantly higher in pre-eclamptic patients when compared with control groups. We concluded that dendritic cells may be involved in the immune regulation during physiological pregnancy. CD1c+ and BDCA-2+ cells can influence the Th2 phenomenon which is observed during physiological pregnancy. Furthermore, it seems possible that lower BDCA-2+ cells percentage and higher CD1c+ : BDCA-2+ ratio can be associated with increased Th1-type immunity in patients with pre-eclampsia.
Increased prevalence of antibodies to enteropathogenicYersinia enterocoliticavirulence proteins in relatives of patients with autoimmune thyroid disease
Authors: STRIEDER T G A , WENZEL B E , PRUMMEL M F , TIJSSEN J G P , WIERSINGA W M
Published: December 2004

Infections have been implicated in the pathogenesis of a number of autoimmune diseases, and Yersinia enterocolitica (YE) might play a role in the development of autoimmune thyroid disease (AITD). Clinical evidence in support of this hypothesis has been inconclusive. We reasoned that looking earlier in the natural course of AITD might enhance chances of finding evidence for YE infection. Consequently, we determined seroreactivity against YE in subjects at risk of developing AITD, i.e. in 803 female relatives of AITD patients in self-proclaimed good health. As a comparison group we used 100 healthy women who participated in a program for reference values. IgG and IgA antibodies to virulence-associated outer membrane proteins (YOPs) of YE were measured by a specific assay. Serum thyroid peroxidase antibodies (TPO-Ab) as indicators of AITD were considered to be positive at levels of> 100 kU/l. The prevalence of YOP IgG-Ab was higher in AITD relatives than in controls (40·1% vs. 24%, P = 0·002), and the same was true for YOP IgA-Ab (22% vs. 13%, P < 0·05). Of the 803 AITD relatives, 44 had an increased or decreased plasma TSH, and 759 were euthyroid as evident from a normal TSH; the prevalence of YOP-Ab did not differ between these three subgroups. TPO-Ab were present in 10% of controls and in 27% of the AITD relatives (P < 0·001). The prevalence of TPO-Ab in the euthyroid AITD relatives was not different between YOP IgG-Ab positive and negative subjects (23·3% vs. 24·7%, NS), nor between YOP IgA-Ab positive and negative subjects (21·2% vs. 24·9%, NS). In conclusion, healthy female relatives of AITD patients have an increased prevalence of YOP antibodies, which, however, is not related to the higher prevalence of TPO antibodies in these subjects. The findings suggest a higher rate of persistent YE infection in AITD relatives. Susceptibility genes for AITD may also confer a risk for YE infection.
Mycophenolic acid inhibits activation of inducible nitric oxide synthase in rodent fibroblasts
Authors: MILJKOVIC Dj , CVETKOVIC I , STOSIC-GRUJICIC S , TRAJKOVIC V
Published: December 2004

Mycophenolate mofetil (MMF) is an immunosuppressive drug that acts as a selective inhibitor of inosine monophosphate dehydrogenase (IMPDH). MMF has recently been shown to inhibit the enzymatic activity of inducible NO synthase (iNOS) and subsequent production of the cytotoxic free radical nitric oxide (NO) in endothelial cells. We here investigated the effect of bioactive MMF compound mycophenolic acid (MPA) on iNOS-mediated NO synthesis in fibroblasts, which are important source of NO in rheumatoid arthritis and during rejection of solid organ transplants. MPA exerted dose-dependent inhibition of NO synthesis, measured as nitrite accumulation, in IFN-γ + LPS-stimulated L929 mouse fibroblast cell line and rat primary fibroblasts. The effect of MPA was not mediated through interference with IMPDH-dependent synthesis of iNOS co-factor BH4 and subsequent suppression of iNOS enzymatic activity, as direct BH4 precursor sepiapterin failed to block the action of the drug. MPA suppressed the IFN-γ + LPS-induced expression of fibroblast iNOS protein, as well as mRNA for iNOS and its transcription factor IRF-1, as assessed by cell-based ELISA and semiquantitative RT-PCR, respectively. MPA suppression of fibroblast NO release, iNOS, and IRF-1 activation, was efficiently prevented by exogenous guanosine, indicating that the drug acted through reduction of IMPDH-dependent synthesis of guanosine nucleotides. These results suggest that MPA inhibits NO production in fibroblasts by blocking guanosine nucleotide-dependent expression of iNOS gene, through mechanisms that might involve the interference with the induction of iNOS transcription factor IRF-1.
Low frequency of plasma nerve-growth factor detection is associated with death of memory B lymphocytes in HIV-1 infection
Authors: TITANJI K , NILSSON A , MÖRCH C , SAMUELSSON A , SÖNNERBORG A , GRUTZMEIER S , ZAZZI M , DE MILITO A
Published: December 2004

Nerve growth factor (NGF) regulates B cell activation and differentiation and is an autocrine survival factor for memory B lymphocytes. We have reported recently that the number of memory B cells is reduced during HIV-1 infection. In this study we evaluated whether alteration in the NGF supply was involved in memory B cell loss in HIV-1-infected subjects. High rate of cell death in vitro was observed in memory B cells from HIV-1-infected individuals compared to uninfected donors (26·2 ± 2·5%versus 7·9 ± 1·4%, P < 0·001). The increased expression of Fas on memory B cells from infected subjects did not enhance the susceptibility of the cells to Fas-mediated apoptosis in vitro. The frequency of NGF detection in plasma from HIV-1-infected subjects was significantly lower than in healthy donors (33·6%versus 63·6%, P < 0·001). Also, the median plasma NGF in HIV-1-infected individuals was significantly lower than in uninfected controls (5 versus 14 pg/ml, respectively, P < 0·01). Interestingly, the plasma NGF level was correlated directly 1to the percentage of memory B cells (P < 0·05). HIV-1-infected subjects with a low number of peripheral memory B cells had a reduced incidence of plasmatic NGF (7·4%) compared to patients with a normal level of memory B cells (37%, P < 0·01). Moreover, the addition of recombinant NGF (1 µg/ml) to cultures of purified B cells reduced cell death of memory B cells from HIV-1-infected subjects from 24·04 ± 3·0% to 17·4 ± 1·3% (P < 0·01). HIV-1-infected individuals also carried higher levels of natural anti-NGF autoantibodies compared to uninfected subjects. In conclusion, we found that memory B cells from HIV-1-infected individuals are primed for cell death. Our study suggests an association between low frequency of plasma NGF detection and the increased cell death of memory B lymphocytes observed during HIV-1 infection. Low levels of NGF in plasma may be due to reduced supply or to NGF binding to natural anti-NGF autoantibodies.
A marked difference in pathogenesis and immune response induced by differentMycobacterium tuberculosisgenotypes
Authors: LÓPEZ B , AGUILAR D , OROZCO H , BURGER M , ESPITIA C , RITACCO V , BARRERA L , KREMER K , HERNANDEZ-PANDO R , HUYGEN K , VAN SOOLINGEN D
Published: December 2004

In the last decade, an unprecedented genetic diversity has been disclosed among Mycobacterium tuberculosis strains found worldwide. However, well-conserved genotypes seem to prevail in areas with high incidence of tuberculosis. As this may be related to selective advantages, such as advanced mechanisms to circumvent [M. bovis Bacille Calmette–Guerin (BCG)-induced] host defence mechanisms, we investigated the influence of strain diversity on the course of experimental disease. Twelve M. tuberculosis strains, representing four major genotype families found worldwide today, and the laboratory strain H37Rv were each used to infect BALB/c mice by direct intratracheal injection. Compared with H37Rv, infections with Beijng strains were characterized by extensive pneumonia, early but ephemeral tumour necrosis factor-alpha (TNF-α) and inducible isoform of nitric oxide synthetase (iNOS) expression, and significantly higher earlier mortality. Conversely, Canetti strains induced limited pneumonia, sustained TNF-α and iNOS expression in lungs, and almost 100% survival. Strains of the Somali and the Haarlem genotype families displayed less homogeneous, intermediate rates of survival. Previous BCG vaccination protected less effectively against infection with Beijing strains than against the H37Rv strain. In conclusion, genetically different M. tuberculosis strains evoked markedly different immunopathological events. Bacteria with the Beijing genotype, highly prevalent in Asia and the former USSR, elicited a non-protective immune response in mice and were the most virulent. Future immunological research, particularly on candidate vaccines, should include a broad spectrum of M. tuberculosis genotypes rather than a few laboratory strains.
Increased susceptibility to airway responses in CD40-deficient mice
Authors: TAKAHASHI H , EBIHARA S , KANDA A , KAMANAKA M , SATO T , HABU S , KIKUTANI H , SASAKI H
Published: December 2004

The interaction between CD40 and its ligand (CD154) is crucial for IL-12 production and effective humoral immunity such as IgE production. Although the interaction seems to play a crucial role in asthmatic inflammation, previous studies investigating the role of the CD40 and CD154 interaction in experimental animal models of asthma are complicated due to multistep reactions in developing asthma. Here, in order to investigate the role of CD40 in the effector phase in the development of airway responses, we used CD40-deficient mice backcrossed with mice transgenic for an ovalbumin (OVA)-specific TCR (TCRtg). Using intranasal OVA administration followed by aerosol inhalation of OVA, greater airway hyperreactivity and eosinophilia in bronchoalveolar lavage fluid (BALF) were observed in CD40-deficient mice backcrossed with TCRtg mice (CD40–/–/ TCRtg mice), compared with control littermates (CD40+/+/ TCRtg mice). CD4+ helper T cell subset analysis of lung draining lymph nodes revealed that the Th1 component was significantly decreased in CD40–/–/ TCRtg mice. Airway hyperreactivity and airway eosinophilia significantly correlated with the predomination of Th2 cells. Cytokine measurements in BALF also showed decreased IL-12 and the predominance of Th2 cells in CD40–/–/ TCRtg mice. These results suggest that CD40 may play a protective role in developing asthma in the phase after establishing specific memory T cells through the regulation of the balance between Th1 and Th2 cells presumably via induction of IL-12.
Suppressive effect of combination treatment of leflunomide and methotrexate on chemokine expression in patients with rheumatoid arthritis
Authors: HO C Y , WONG C K , LI E K , TAM L S , LAM C W K
Published: December 2004

To study the immunosuppressive and anti-inflammatory effects of combined leflunomide and methotrexate (MTX) therapy on chemokine expression in patients with rheumatoid arthritis (RA), nine patients were enrolled for the combination therapy for 24 weeks. These patients have been on treatment with MTX 15 mg/week for not less than 3 months before entry to the study. A loading dose of l00 mg/day of leflunomide was given for 3 days, followed by 10 mg/day for the rest of the study period. Plasma concentrations of monocyte chemotactic protein-1 (MCP-1), thymus- and activation-regulated chemokine (TARC), and macrophage-derived chemokine (MDC) were assayed before and after combination treatment by ELISA. Gene expression of inflammatory cytokines and chemokines of peripheral blood mononuclear cells was analysed by cDNA expression array. Plasma MCP-1, TARC and MDC concentrations were significantly lower in patients after combination treatment [median (interquartile range) before versus after treatment: MCP-1 of 118·0 (64·0–515·2) versus 3·2 (0·0–22·8) pg/ml, P < 0·01; TARC of 126·1 (27·2–197·4) versus 0·0 (0·0–52·5) pg/ml, P < 0·05; MDC of 503·3 (446·2–600·9) versus 366·8 (337·4–393·4) pg/ml, P < 0·05]. Positive correlations among reductions in plasma chemokines and clinical outcome measures were also found. Expression of chemokine genes including MDC and TARC was suppressed after combination treatment [% suppression of 38·7 (54·3–13·0) and 53·7 (55·9–28·4), respectively]. Combination therapy with leflunomide and MTX exhibits anti-inflammatory activity in the suppression of chemokine expression and subsequent recruitment of inflammatory cells into the inflammatory sites in RA.