bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of human leucocyte antigen mismatch on graft‐versus‐host disease and graft failure after reduced intensity conditioning allogeneic haematopoietic stem cell transplantation from related donors
Authors: Teshima Takanori , Matsuo Keitaro , Matsue Kosei , Kawano Fumio , Taniguchi Shuichi , Hara Masamichi , Hatanaka Kazuo , Tanimoto Mitsune , Harada Mine , Nakao Shinji , Abe Yasunobu , Wake Atsushi , Eto Tetsuya , Takemoto Yoshinobu , Imamura Masahiro , Takahashi Satoshi , Ishida Yoji , Kanda Yoshinobu , Kasai Masaharu , Takaue Yoichi
Published: July 2005

The impact of human leucocyte antigen (HLA) incompatibility between donor and recipient on graft‐versus‐host disease (GVHD) and graft failure after reduced‐intensity conditioning stem cell transplantation (RICT) remains to be elucidated. We retrospectively analysed outcome in 341 patients who underwent RICT from related donors for haematological malignancies. The overall cumulative incidence of grade II–IV acute GVHD (aGVHD) was 40% for all subjects; 39% in recipients with HLA‐matched donors, 44% in those with one‐locus‐mismatched donors, and 50% in those with two‐ to three‐loci‐mismatched donors. In a Cox regression model adjusted for potential confounders, the tendency for grade II–IV aGVHD (P = 0·01), chronic GVHD (cGVHD) (P = 0·05) and graft failure (P = 0·033) increased with HLA disparity. Use of peripheral blood grafts instead of marrow was a risk factor for cGVHD. Use of antithymocyte globulin was associated with reduced aGVHD and cGVHD. Overall survival (OS) in recipients of two‐ to three‐loci‐mismatched RICT at 2 years (18%) was significantly worse than that in patients who received one‐locus‐mismatched RICT (51%) and HLA‐matched RICT (48%) (P < 0·0001). A two‐ to three‐loci mismatch was identified as an independent risk factor for OS (P < 0·001), but there was no significant difference in OS between HLA‐matched and one‐locus‐mismatched RICT. HLA incompatibility between the donor and recipient is an important risk factor for graft failure, aGVHD, cGVHD and OS after RICT. RICT from a one‐locus‐mismatched donor may represent an effective alternative approach in patients with high‐risk malignancies who lack HLA‐matched related donors.
Immunological profile of peripheral blood lymphocytes and monocytes/macrophages in Kawasaki disease
Authors: Matsubara T , Ichiyama T , Furukawa S
Published: July 2005

Kawasaki disease (KD) is an acute illness of early childhood characterized by prolonged fever, diffuse mucosal inflammation, indurative oedema of the hands and feet, a polymorphous skin rash and nonsuppurative lymphadenopathy. The histopathological findings in KD comprise panvasculitis with endothelial necrosis, and the infiltration of mononuclear cells into small and medium-sized blood vessels. The levels of many proinflammatory cytokines, chemokines and adhesion molecules can be elevated in sera from children with KD at the acute stage. Although many immunological studies on KD involving peripheral blood have been reported, the data obtained remain controversial. This review focuses on the immune response of peripheral blood lymphocytes and monocytes/macrophages during acute KD.
Insights into the role of fibroblasts in human autoimmune diseases
Authors: Smith T J
Published: July 2005

Traditional wisdom has considered fibroblasts as contributing to the structural integrity of tissues rather than playing a dynamic role in physiological or pathological processes. It is only recently that they have been recognized as comprising diverse populations of cells exhibiting complex patterns of biosynthetic activity. They represent determinants that react to stimuli and help define tissue remodelling through the expression of molecules imposing constraints on their cellular neighbourhood. Moreover, fibroblasts can initiate the earliest molecular events leading to inflammatory responses. Thus they must now be viewed as active participants in tissue reactivity. In this short review, I will provide an overview of contemporary thought about the contribution of fibroblasts to the pathogenesis of autoimmune processes through their expression of, and responses to, mediators of inflammation and tissue remodelling.
CXCL12 and CXCR4 expression by human gingival fibroblasts in periodontal disease
Authors: Hosokawa Y , Hosokawa I , Ozaki K , Nakae H , Murakami K , Miyake Y , Matsuo T
Published: July 2005

CXCL12 is a CXC chemokine that is related to lymphocyte infiltration and angiogenesis in inflammatory sites such as arthritis. However, the expression and roles of CXCL12 in periodontal disease are uncertain. The aim of this study was to assess the expression of CXCL12 and its receptor, CXCR4, in periodontal tissue and to investigate the properties of CXCL12 and CXCR4 expression by human gingival fibroblasts (HGF). RT-PCR analysis revealed that CXCL12 and CXCR4 mRNA were expressed in both normal gingival tissues and periodontal diseased tissues. Immunohistochemistry disclosed that CXCL12 was expressed and CXCR4 positive cells were found in both normal and periodontal diseased gingival tissues. Our in vitro experiments elucidated that HGF constitutively produced CXCL12, and the levels were enhanced by stimulation with tumour necrosis factor-α (TNF-α), interferon-γ (IFN-γ), transforming growth factor-β (TGF–β), regulated upon activation normal T cell expressed and secreted (RANTES) and macrophage inflammatory protein 3α (MIP-3α). On the other hand, heat killed Porphyromonas gingivalis (P. gingivalis) and P. gingivalis LPS reduced the CXCL12 production by HGF. Flow cytometry analysis clarified that CXCR4 was highly expressed on HGF, and CXCR4 expression was abrogated by TNF-α, IFN-γ and P. gingivalis LPS. Moreover, CXCL12 induced vascular endothelial growth factor (VEGF) production by HGF. Our results demonstrated that CXCL12 might be related to CXCR4+ cells infiltration and angiogenesis both in normal periodontal tissues and periodontal diseased tissue. P. gingivalis, a known periodontal pathogen, inhibits the production of CXCL12 and the expression of CXCR4 by HGF. This fact means that P. gingivalis may inhibit CXCR4+ cells infiltration and neovascularization in periodontal tissue and escape from the immune response.
Toxoplasma gondiiregulates recruitment and migration of human dendritic cells via different soluble secreted factors
Authors: Diana J , Vincent C , Peyron F , Picot S , Schmitt D , Persat F
Published: July 2005

We investigated in vitro the properties of soluble factors produced by Toxoplasma gondii on the recruitment, maturation and migration of human dendritic cells (DC) derived from CD34+ progenitor cells. We used soluble factors including excreted secreted antigens (ESA) produced under various conditions by the virulent type I RH strain (ESA-RH) and the less virulent PRU type II strain (ESA-PRU). Soluble factors of both T. gondii strains appeared to possess a chemokine-like activity that attracted immature DC. This recruitment activity required the presence of functional CCR5 molecules on the cell membrane. Incubation of DC for 24 h with ESA triggered the migration of a large percentage of these cells towards the chemokine MIP-3β; ESA-PRU was more efficient than ESA-RH. ESA produced in absence of exogenous protein and crude extract did not induce DC migration but retained recruitment activity. These data indicate that recruitment activity and migration-inducing activity are not governed by the same factors. Moreover, incubation of DC for 48 h with ESA did not modify the expression of costimulation or maturation markers (CD83, CD40, CD80, CD86 or HLA-DR), but induced a decrease in CCR6 expression associated with an increased expression of CCR7. Taken together, these results suggest that T. gondii controls recruitment and migration of immature DC by different soluble factors and may induce a dysfunction in the host-specific immune response.
CDC staging based on absolute CD4 count and CD4 percentage in an HIV-1-infected Indian population: treatment implications
Authors: Vajpayee M , Kaushik S , Sreenivas V , Wig N , Seth P
Published: July 2005

CD4+ T-cell levels are an important criterion for categorizing HIV-related clinical conditions according to the CDC classification system and are therefore important in the management of HIV by initiating antiretroviral therapy and prophylaxis for opportunistic infections due to HIV among HIV-infected individuals. However, it has been observed that the CD4 counts are affected by the geographical location, race, ethnic origin, age, gender and changes in total and differential leucocyte counts. In the light of this knowledge, we classified 600 HIV seropositive antiretroviral treatment (ART)-naïve Indian individuals belonging to different CDC groups A, B and C on the basis of CDC criteria of both CD4% and CD4 counts and receiver operating characteristic (ROC) curves were generated. Importantly, CDC staging on the basis of CD4% indicated significant clinical implications, requiring an early implementation of effective antiretroviral treatment regimen in HIV-infected individuals deprived of treatment when classified on the basis of CD4 counts.
HUVECs from newborns with a strong family history of myocardial infarction overexpress adhesion molecules and react abnormally to stimulating agents
Authors: Paez A , Méndez-Cruz A R , Varela E , Rodriguez E , Guevara J , Flores-Romo L , Montaño L F , Massó F A
Published: July 2005

Atherosclerosis is a complex disease involved in major fatal events such as myocardial infarction and stroke. It is the result of interactions between metabolic, dietetic and environmental risk factors acting on a genetic background that could result in endothelial susceptibility. Our aim was to determine the patterns of expression of adhesion molecules and whether phosphatidylserine is translocated to the cell surface of human umbilical vein endothelial cells (HUVECs) isolated from healthy newborns born to parents with a strong family history of myocardial infarction under TNF-α or oxLDL stimulated conditions. Compared to control HUVECs, experimental cords showed: (a) a four-fold increase in VCAM-1 expression under basal conditions, which showed no change after stimulation with the pro-atherogenic factors; (b) a two-fold increase in basal P-selectin expression that reached a 10-fold increase with any of the pro-atherogenic factors; (c) a basal ICAM-1 expression similar to P-selectin that was not modified by the pro-atherogenic molecules; (d) a similar PECAM-1 expression. Unexpectedly, phospathidylserine expression in experimental cord HUVECs was significantly increased (211 817 versus 3354 TFU) but was not associated to apoptotic death as the percentage of dead cells induced by TNF-α treatment was very low (0·55 versus 9·87% in control HUVECs). The latter result was corroborated by TUNEL staining. T cell adherence to HUVECs was highly up-regulated in the genetically predisposed samples. The analysis of nonpooled HUVECs, from newborns to family predisposed myocardial-infarction individuals, might represent a useful strategy to identify phenotypical and functional alterations, and hopefully, to take early preventive actions.
Circulating anti-actin and anti-ATP synthase antibodies identify a sub-set of patients with idiopathic nephrotic syndrome
Authors: Musante L , Candiano G , Bruschi M , Santucci L , Carnemolla B , Orecchia P , Giampuzzi M , Zennaro C , Sanna-Cherchi S , Carraro M , Oleggini R , Camussi G , Perfumo F , Ghiggeri G M
Published: July 2005

Idiopathic nephrotic syndrome (iNS) with resistance or dependence to steroids is a common disease in children but in spite of an increasing clinical impact its pathogenesis is unknown. We screened for the presence of circulating antibodies against glomerular (podocytes, mesangium) and tubular cells (tubular epithelia) a cohort of 60 children with iNS including 8 patients with a familial trait of iNS or with proven mutation of NPHS1-NPHS2 and 12 with good sensitivity to steroids. Positive sera were found in 8 cases, all belonging to the category without familial trait/molecular defects. The targets of antibodies were characterized with Western blot and MALDI-Mass utilizing β-hexyl cell extracts separated with two-dimensional electrophoresis. In all cases antibodies of the IgM class were directed against ATP synthase β chain alone (4 cases) or in combination with actin (3 cases); one child presented IgG against aldose reductase. The clinical picture was nephrotic syndrome with steroid resistance or dependence and variable cyclosporin sensitivity; 3 patients developed end stage renal failure. The basic pathology picture was focal segmental glomerulosclerosis (FSGS) in 4 cases and mesangial proliferative glomerulonephrites with deposition of IgM in 2. Overall, patients with circulating auto-antibodies could not be readely differentiated on clinical grounds with the exception of 3 children who developed positivity for antinuclear antibodies during the follow-up. Affinity-purified IgM from one patient who underwent plasmapheresis for therapeutical pourposes (but not from a normal pool) induced proteinuria in Sprague-Dawley rats and concomitant human IgM deposition within glomeruli. This is the first report of circulating anti-actin/ATP synthase β chain antibodies in a subset of patients with iNS. Both pathological significance and clinical impact given by the presence of these antibodies and the relationship with other conditions such as lupus-erythematosus, characterized by their presence, must be defined.
Ex vivostimulation of cord blood mononuclear cells by dexamethasone and interleukin-7 results in the maturation of interferon-γ-secreting effector memory T cells
Authors: Talayev V Yu , Zaichenko I Ye , Babaykina O N , Lomunova M A , Talayeva E B , Nikonova M F
Published: July 2005

The effects of dexamethasone phosphate and interleukin-7 upon the proliferation of T-cells and the production of interferon-γ in the newborn's cord blood mononuclear cell cultures were studied. The capability of dexamethasone to enhance T-cell proliferation induced by anti-CD3 with interleukin-7 in some newborn cord blood mononuclear cell cultures was identified. Dexamethasone suppressed production of interferon-γ in 68-h cell cultures stimulated with anti-CD3 both in the presence of interleukin-7 and without it. However, a 68-h cultivation of newborn blood cells with dexamethasone, anti-CD3 and interleukin-7 resulted in the accumulation of T-lymphocytes capable of producing interferon-γ after restimulation. As a result of it the amount of interferon-γ producing CD7+ T-cells and the concentration of interferon-γ in cultural supernatants were maximal in the cell cultures incubated with anti-CD3, interleukin-7 and dexamethasone during the first 68 h and subsequently restimulated with phorbol 12-myristate 13-acetate and ionomycin. The stimulation of neonatal or adult blood cells by dexamethasone, anti-CD3 and interleukine-7 also causes a decrease in the number of naïve T-cells and central memory cells and an increase in the number of effector memory CD7+CD45RA+CD62L– cells in cultures. It is possible that these effects are caused by the influence of dexamethasone on IL-7 receptor expression: it is known that IL-7 receptor alpha-chain gene is a glucocorticoid-inducible gene.
Antibody to severe acute respiratory syndrome (SARS)-associated coronavirus spike protein domain 2 cross-reacts with lung epithelial cells and causes cytotoxicity
Authors: Lin Y S , Lin C F , Fang Y T , Kuo Y M , Liao P C , Yeh T M , Hwa K Y , Shieh C C K , Yen J H , Wang H J , Su I J , Lei H Y
Published: July 2005

Both viral effect and immune-mediated mechanism are involved in the pathogenesis of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) infection. In this study, we showed that in SARS patient sera there were autoantibodies (autoAbs) that reacted with A549 cells, the type-2 pneumocytes, and that these autoAbs were mainly IgG. The autoAbs were detectable 20 days after fever onset. Tests of non-SARS-pneumonia patients did not show the same autoAb production as in SARS patients. After sera IgG bound to A549 cells, cytotoxicity was induced. Cell cytotoxicity and the anti-epithelial cell IgG level were positively correlated. Preabsorption and binding assays indicated the existence of cross-reactive epitopes on SARS-CoV spike protein domain 2 (S2). Furthermore, treatment of A549 cells with anti-S2 Abs and IFN-γ resulted in an increase in the adherence of human peripheral blood mononuclear cells to these epithelial cells. Taken together, we have demonstrated that the anti-S2 Abs in SARS patient sera cause cytotoxic injury as well as enhance immune cell adhesion to epithelial cells. The onset of autoimmune responses in SARS-CoV infection may be implicated in SARS pathogenesis.
An increased MRP8/14 expression and adhesion, but a decreased migration towards proinflammatory chemokines of type 1 diabetes monocytes
Authors: Bouma G , Coppens J M C , Lam-Tse W-K , Luini W , Sintnicolaas K , Levering W H , Sozzani S , Drexhage H A , Versnel M A
Published: July 2005

In the early development of type 1 diabetes macrophages and dendritic cells accumulate around the islets of Langerhans at sites of fibronectin expression. It is thought that these macrophages and dendritic cells are derived from blood monocytes. Previously, we showed an increased serum level of MRP8/14 in type 1 diabetes patients that induced healthy monocytes to adhere more strongly to fibronectin (FN). Here we show that MRP8/14 is expressed and produced at a higher level by type 1 diabetes monocytes, particularly after adhesion to FN, creating a positive feedback mechanism for a high fibronectin-adhesive capacity. Also adhesion to endothelial cells was increased in type 1 diabetes monocytes. Despite this increased adhesion the transendothelial migration of monocytes of type 1 diabetes patients was decreased towards the proinflammatory chemokines CCL2 and CCL3. Because non-obese diabetic (NOD) mouse monocytes show a similar defective proinflammatory migration, we argue that an impaired monocyte migration towards proinflammatory chemokines might be a hallmark of autoimmune diabetes. This hampered monocyte response to proinflammatory chemokines questions whether the early macrophage and dendritic cell accumulation in the diabetic pancreas originates from an inflammatory-driven influx of monocytes. We also show that the migration of type 1 diabetes monocytes towards the lymphoid tissue-related CCL19 was increased and correlated with an increased CCR7 surface expression on the monocytes. Because NOD mice show a high expression of these lymphoid tissue-related chemokines in the early pancreas it is more likely that the early macrophage and dendritic cell accumulation in the diabetic pancreas is related to an aberrant high expression of lymphoid tissue-related chemokines in the pancreas.
Sam68 is tyrosine phosphorylated and recruited to signalling in peripheral blood mononuclear cells from HIV infected patients
Authors: Najib S , Rodríguez-Baño J , Ríos M J , Muniain M A , Goberna R , Sánchez-Margalet V
Published: July 2005

Human immunodeficiency virus (HIV) codes for a protein, Rev, that mediates the viral RNA export from the nucleus to the cytoplasm. Recently, it has been found that Sam68, the substrate of Src associated in mitosis, is a functional homologue of Rev, and a synergistic activator of Rev activity. Thus, it has been suggested that Sam68 may play an important role in the post-transcriptional regulation of HIV. Sam68 contains an RNA binding motif named KH [homology to the nuclear ribonucleoprotein (hnRNP) K]. Tyrosine phosphorylation of Sam68 and binding to SH3 domains have been found to negatively regulate its RNA binding capacity. Besides, tyrosine phosphorylation of Sam68 allows the formation of signalling complexes with other proteins containing SH2 and SH3 domains, suggesting a role in signal transduction of different systems in human lymphocytes, such as the T cell receptor, and leptin receptor, or the insulin receptor in other cell types. In the present work, we have found that Sam68 is tyrosine phosphorylated in peripheral blood mononuclear cells (PBMC) from HIV infected subjects, leading to the formation of signalling complexes with p85 the regulatory subunit of PI3K, GAP and STAT-3, and decreasing its RNA binding capacity. In contrast, PBMC from HIV infected subjects have lower expression levels of Sam68 compared with controls. These results suggest that Sam68 may play some role in the immune function of lymphocytes in HIV infection.
Histological analysis of CD11c-DTR/GFP mice after in vivo depletion of dendritic cells
Authors: Probst H C , Tschannen K , Odermatt B , Schwendener R , Zinkernagel R M , Van Den Broek M
Published: July 2005

To investigate the dependence of individual immunological processes on DC, a transgenic mouse system (CD11c-DTR/GFP mice) has been developed that allows conditional depletion of CD11c+ DC in vivo through administration of diphtheria toxin. We have performed careful histological analysis of CD11c-DTR/GFP mice at different time points after diphtheria toxin injection and confirmed the transient depletion of CD11c+ cells from lymph nodes and spleen. Unexpectedly, the injection of diphtheria toxin completely depleted marginal zone and metallophilic MΦ from the spleen and their sinusoidal counterparts from the lymph nodes. This finding limits the use of CD11c-DTR/GFP mice for the analysis of the role of DC to models and read outs that are proven to be independent of marginal zone and sinusoidal MΦ.
2B4 expression on natural killer cells increases in HIV-1 infected patients followed prospectively during highly active antiretroviral therapy
Authors: Ostrowski S R , Ullum H , Pedersen B K , Gerstoft J , Katzenstein T L
Published: July 2005

Human immunodeficiency virus (HIV)-1 infection influences natural killer (NK) cell expression of inhibitory NK receptors and activating natural cytotoxicity receptors. It is unknown whether expression of the co-stimulatory NK cell receptor 2B4 (CD244) on NK cells and CD3+ CD8+ cells are affected by highly active antiretroviral therapy (HAART), low-level viraemia, proviral-DNA or immune activation in HIV-1 infected patients. A total of 101 HAART-treated HIV-1 infected patients with ≤ 200 HIV-RNA copies/ml were followed prospectively for 24 months. HIV-RNA was investigated 3-monthly and 2B4 expression on CD3– CD16+ NK cells and CD3+ CD8+ cells, proviral-DNA and plasma soluble tumour necrosis factor receptor (sTNFr)-II were investigated 6-monthly. For comparison, 2B4 expression was investigated in 20 healthy individuals. The concentration of 2B4+ NK cells was initially reduced in HIV-1 infected patients (P < 0·001) but increased to a normal level during the 24 months’ follow-up. The concentration of CD3+ CD8+ 2B4+ cells in HIV-1 infected patients was normal and did not change during follow-up. The relative fluorescence intensity (RFI) of 2B4 increased on both NK cells and CD3+ CD8+ cells during follow-up (both P < 0·001). Higher levels of proviral-DNA carrying cells and plasma sTNFrII were associated with reductions in the concentration of 2B4+ NK cells (all P < 0·05). HIV-RNA had no effect on 2B4 expression on NK cells or CD3+ CD8+ cells. These findings demonstrate that the concentration of 2B4+ NK cells normalizes during long-term HAART in HIV-1 infected patients. The finding that proviral-DNA and sTNFrII were associated negatively with the concentration of 2B4+ NK cells suggests that immune activation in HIV-1 infected patients receiving HAART influences the target cell recognition by NK cells.
The direct and indirect allogeneic presentation pathway during acute rejection after human cardiac transplantation
Authors: van Besouw N M , Zuijderwijk J M , Vaessen L M B , Balk A H M M , Maat A P W M , van der Meide P H , Weimar W
Published: July 2005

Alloreactive T cells may be activated via a direct or an indirect antigen presentation pathway. We questioned whether the frequency of interferon (IFN)-γ producing cells determined by enzyme-linked immunospot (ELISPOT) assay is an effective tool to monitor the direct and/or indirect presentation pathway. Secondly, we wondered whether early and late acute rejection (AR) are associated with both pathways. Before (n = 15), during (n = 18) and after (n = 16) a period of AR, peripheral blood mononuclear cell (PBMC) samples were tested from 13 heart transplant recipients. The direct presentation pathway was always present. The number of IFN-γ producing cells reactive to this pathway increased significantly (P = 0·04) during AR and the number decreased (P = 0·005) after AR therapy. In contrast, the indirect allogeneic presentation pathway was present in only eight of 18 AR samples. When the indirect presentation pathway was detectable, it increased significantly during AR. Five of eight of these AR occurred more than 6 months after transplantation. The ELISPOT assay, enumerating alloreactive IFN-γ producing cells, is a valuable tool to determine the reactivity via both the direct and the indirect presentation pathway. The direct presentation pathway always plays a role in AR, while the indirect pathway contributes especially to late AR.
Detection ofin vitrointerferon-γ and serum tumour necrosis factor-α in multidrug-resistant tuberculosis patients
Authors: Fortes A , Pereira K , Antas P R Z , Franken C L M C , Dalcolmo M , Ribeiro-Carvalho M M , Cunha K S , Geluk A , Kritski A , Kolk A , Klatser P , Sarno E N , Ottenhoff T H M , Sampaio E P
Published: July 2005

Multidrug-resistant tuberculosis (MDR-TB) is known as having a poor prognosis with a weak response to therapy and very high death rates. The aim of this work was to assess the immune response to the RD1-encoded antigen ESAT-6 of Mycobacterium tuberculosis in MDR-TB patients and compare to non-resistant (NR) TB patients and healthy controls (HC). Evaluation of interferon (IFN)-γ production showed that, although 55% of the MDR patients were responsive to ESAT-6, they produced lower IFN-γ levels (553 ± 11 pg/ml) when compared to NR-TB (1179 ± 163 pg/ml; P < 0·05) but not to controls (412 ± 65·7 pg/ml). Differences in the response to ESAT-6 and to its overlapping peptides mixture were also significant between MDR versus treated pulmonary NR-TB. Furthermore, a very low rate of response to PPD (23·5%) and to Ag85B (33·3%) was noted in MDR-TB patients as compared to the other groups. To determine the inflammatory response in patients’ groups, detection of tumour necrosis factor (TNF)-α was assessed in their sera before and during chemotherapy. Mean TNF-α levels in MDR-TB (43·8 ± 9 pg/ml) paralleled those found in treated pulmonary, and it was significantly different (P < 0·05) from the values found in untreated NR and HC. Interestingly, secretion of IFN-γ and TNF-α were predominant in MDR patients who presented with bilateral pulmonary lesions and lung cavitation. The present data indicate that the overall immune response to mycobacterial antigens is decreased in resistant TB and the major role inflammatory cytokines may play in perpetuating pulmonary tissue damage.
Adenovirus-mediated gene delivery of interleukin-10, but not transforming growth factor β, ameliorates the induction of Graves’ hyperthyroidism in BALB/c mice
Authors: Saitoh O , Mizutori Y , Takamura N , Yamasaki H , Kita A , Kuwahara H , Nagayama Y
Published: July 2005

Interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) are well known anti-inflammatory cytokines. We have studied the effect of adenovirus-mediated IL-10 and TGF-β gene delivery on the induction of Graves’ hyperthyroidism in our mouse model that involves repeated injections of adenovirus expressing the thyrotropin receptor A subunit (AdTSHR). We first constructed adenoviruses encoding the two cytokines (AdIL10 and AdTGFβ) and confirmed expression by in vitro infection of COS cells. Susceptible BALB/c mice were injected twice with AdTSHR alone or together with AdIL10 or AdTGFβ, and bled two weeks after the second immunization. Significantly elevated serum thyroxine levels were seen in 26% of mice immunized with AdTSHR and AdIL10 versus 61% with AdTSHR alone. Levels of thyroid stimulating antibody, but not nonstimulating antibody, were also decreased, and TSHR-specific splenocyte secretion of interferon-γ in recall assays was impaired in mice treated with AdIL10. In contrast, AdTGFβ had little effect on hyperthyroidism. Overall, our findings demonstrate that gene delivery of IL-10, but not TGF-β, suppresses the induction of Graves’ hyperthyroidism in a mouse model. However, the effect of IL-10 is less powerful than we observed previously with T helper type 2-inducers including adenovirus expressing IL-4, Shistosoma mansoni infection or α-galactosylceramide.
Interleukin-4 and interleukin-13 enhance CCL26 production in a human keratinocyte cell line, HaCaT cells
Authors: Kagami S , Saeki H , Komine M , Kakinuma T , Tsunemi Y , Nakamura K , Sasaki K , Asahina A , Tamaki K
Published: July 2005

Eotaxin-2/CCL24 and eotaxin-3/CCL26 are CC chemokines and their receptor, CC chemokine receptor 3 is preferentially expressed on eosinophils. It was reported that vascular endothelial cells and dermal fibroblasts produced CCL26. However, the regulation of CCL24 and CCL26 production in keratinocytes has not been well documented. We investigated the expression and production of CCL24 and CCL26 in the human keratinocyte cell line, HaCaT cells. Reverse transcription and polymerase chain reaction was performed using these cells and Enzyme-linked immunosorbent assay was carried out using supernatant of these cells. The production of CCL24 in HaCaT cells was slightly enhanced by IL-4 and that of CCL26 was strongly enhanced by IL-4 and IL-13. Furthermore, TNF-α generated a synergistic effect on IL-4 enhanced CCL26 production. Dexamethasone, IFN-γ and the p38 mitogen-activated protein kinase inhibitor SB202190 inhibited IL-4 enhanced CCL26 production. IL-4 enhanced production of CCL26 was inhibited by leflunomide and JAK inhibitor 1, but not by JAK3 inhibitor, which indicates that it is mediated by JAK1-STAT6-dependent pathway. This result also strongly suggests the involvement of the type 2 IL-4 receptor in IL-4 enhanced production of CCL26. These results suggest that keratinocytes are involved in the migration of CC chemokine receptor 3 positive cells such as eosinophils in a Th2-dominant situation like atopic dermatitis.
FoxP3+CD4+CD25+ T cells with regulatory properties can be cultured from colonic mucosa of patients with Crohn's disease
Authors: Kelsen J , Agnholt J , Hoffmann H J , Rømer J L , Hvas C L , Dahlerup J F
Published: July 2005

CD4+CD25+ regulatory T cells (Tregs) are involved in the maintenance of peripheral tolerance and ensure a balanced immune response competent of fighting pathogens and at the same time recognizing commensals as harmless. This feature is lost in Crohn's disease (CD). The forkhead/winged helix transcription factor FoxP3 is a master gene for Treg function and defects in the FoxP3 gene lead to a clinical picture similar to inflammatory bowel disease (IBD). Murine colitis can be cured by adoptive transfer of Tregs and ex vivo-generated gut-specific Tregs represent an attractive option for therapy in CD. Thus, defective Tregs could contribute to the development of CD. We cultured biopsies of colonic mucosa in the presence of high concentrations of interleukin (IL)-2 and IL-4 to overcome the anergic nature of naturally occurring CD4+CD25+ Tregs in the mucosa. We investigated the expression of FoxP3 and regulatory potential of gut-derived CD4+CD25+ T cells cultured from patients with CD and healthy individuals. The FoxP3 expression was analysed by reverse transcriptase polymerase chain reaction (RT-PCR), and the suppressive effect of FoxP3+CD4+CD25+ T cells on proliferation and cytokine production of autologous CD4+ T cells was assessed by flow cytometry. Cultured gut-derived T cells with CD4+CD25+ phenotype expressed FoxP3 and were able as the freshly isolated Tregs from peripheral blood to suppress proliferation and cytokine production of autologous CD4+ T cells. Thus, we demonstrate that FoxP3+CD4+CD25+ T cells with regulatory properties can be propagated in vitro from inflamed mucosa of CD patients, which may be of interest in adoptive immunotherapy.
Macrophage migration inhibitory factor contributes to the development of acute dextran sulphate sodium-induced colitis in Toll-like receptor 4 knockout mice
Authors: Ohkawara T , Takeda H , Nishihira J , Miyashita K , Nihiwaki M , Ishiguro Y , Takeda K , Akira S , Iwanaga T , Sugiyama T , Asaka M
Published: July 2005

Toll-like receptor 4 (TLR4), which recognizes lipopolysaccharides, plays an important role in the innate immune response. In this study, we investigated the role of TLR4 in the development of experimental colitis with regard to the biological actions of macrophage migration inhibitory factor (MIF) using TLR4 null (–/–) mice. TLR4–/– mice were given 2% dextran sulphate sodium (DSS) in drinking water to induce colitis, which was clinically and histologically as severe as that seen in wild-type (WT) mice. The level of tumour necrosis factor (TNF)-α in colon tissues was increased in WT mice but unchanged in TLR4–/– mice. The level of myeloperoxidase (MPO) activity in colon tissues was increased by DSS administration in both TLR4–/– and WT mice. The expression of MIF was up-regulated in the colons of TLR4–/– mice with acute DSS-induced colitis. An anti-MIF antibody significantly suppressed colitis and elevation of matrix metalloproteinase-13 in TLR4–/– mice. The current results obtained from TLR4–/– mice provide evidence that MIF plays a critical role in the development of acute DSS-induced colitis.
Interleukin-10 in murine metal-induced systemic autoimmunity
Authors: Häggqvist B , Hultman P
Published: July 2005

Systemic autoimmune diseases have a complicated and largely unknown aetiology and pathogenesis, but they are at least partly obeying the rules of an ordinary immune response. Cytokines are therefore important in the pathogenesis as demonstrated by the recent success in treating rheumatoid arthritis with anti-cytokine agents. The suppressive functions in the immune system have lately received much interest. One of the cytokines in focus in this respect is interleukin (IL)-10. We recently observed that in heavy-metal induced systemic autoimmunity, genetically resistant mice show a strong increase in IL-10 mRNA expression, which was not seen in susceptible mice. We have therefore examined the possible regulating effect of IL-10 on the induction and manifestation of systemic autoimmunity in this model. We took two approaches: a targeted mutation of the IL-10 gene in a strain resistant to heavy metal-induced autoimmunity, and treatment with recombinant IL-10 in the genetically susceptible A.SW strain during the induction of autoimmunity by metals. The wild-type C57BL/6 J (B6-WT) strain did not react with lymphoproliferation, polyclonal B cell activation, anti-nucleoar autoantibodies (ANoA) or tissue immune-complex (IC) deposits in response to inorganic mercury (Hg) or silver (Ag). However, serum IgG1 and IgE showed a modest increase during Hg treatment, while Ag caused a weak increase in IgE and IgG2a. The B6·129P2-Il10tm1Cgn/J strain (IL-10-deficient mice) did not develop antinucleolar antibodies (ANoA) during Hg treatment, but showed a higher median titre of homogeneous ANA compared with Hg-treated B6-WT mice. Both control and Hg-treated (but not Ag-treated) IL-10-deficient mice showed an increase in splenic weight and serum IgG1 compared with B6-WT control and Hg-treated mice. An early, significant increase in serum IgE was seen in Hg-treated IL-10-deficient and WT mice compared with the controls; the increase was 42- and sixfold, respectively. During ongoing intense treatment with rIL-10 in combination with Hg the susceptible A.SW mice showed a reduced development of ANoA and antichromatin antibodies, as well as serum IgE, compared with mice receiving Hg but not rIL-10. In conclusion, IL-10 suppresses several aspects of HgIA, but is not crucial for resistance to heavy metal-induced autoimmunity. Peroral silver treatment suppresses the spontaneous immune activation seen in IL-10-deficient mice.
Rare naturally occurring immune responses to three epitopes from the widely expressed tumour antigens hTERT and CYP1B1 in multiple myeloma patients
Authors: Maecker B , Von Bergwelt-Baildon M S , Anderson K S , Vonderheide R H , Anderson K C , Nadler L M , Schultze J L
Published: July 2005

The widely expressed tumour antigens hTERT and CYP1B1 are commonly expressed in multiple myeloma (MM) cells. Several trials targeting these antigens by immunotherapy have been initiated. The aim of this study was to explore whether patients with MM have an endogenous pre-existing immune response against recently identified epitopes from hTERT and CYP1B1. Peripheral blood T cells from 27 HLA-A*0201+ multiple myeloma patients at different stages of disease and 20 healthy HLA-A*0201+ donors were enriched and studied for the presence of hTERT- and CYP1B1-specific cytotoxic T cells using MHC tetramer detection and short-term ex vivo expansion. No significant expansion of tetramer-positive cells was detected in the peripheral blood of either MM patients or healthy controls when cells were stained with tetramers containing the dominant hTERT-derived epitope or two peptides derived from CYP1B1. A single ex vivo peptide stimulation led to the detection of a small population (0·3–0·5%) of hTERT-specific cells in two of 27 patients with MM. None of the patients or controls showed significant expansion of CYP1B1-specific cells after a single peptide stimulation. Thus, endogenous in vivo priming of T cells against hTERT and CYP1B1 is a rare event in MM patients. These results suggest that strategies targeting hTERT and CYP1B1 may have to utilize techniques to induce T cell responses from a naive precursor frequency.
Exposure toMycobacterium aviuminduces low-level protection fromMycobacterium bovisinfection but compromises diagnosis of disease in cattle
Authors: Hope J C , Thom M L , Villarreal-Ramos B , Vordermeier H M , Hewinson R G , Howard C J
Published: July 2005

We assessed the effect of exposure to Mycobacterium avium on the development of immune responses and the pathogenesis of disease observed following Mycobacterium bovis challenge. A degree of protection against M. bovis was observed in calves which were pre-exposed to M. avium as assessed by the extent of lesions and bacterial load compared to the M. bovis alone group. The immune response following M. bovis challenge in cattle previously inoculated with M. avium was biased towards antigens (PPD) present in M. avium, whereas the response following M. bovis alone was biased towards antigens present in M. bovis, indicating an imprinting of memory to avian antigens on T lymphocytes. A consequence of the memory to M. avium antigens was failure to diagnose M. bovis infection by the skin test or the IFNγ assay in some of the animals which had lesions of tuberculosis at necropsy. The use of M. bovis specific antigens ESAT-6 and CFP-10 increased IFNγ test specificity in animals previously exposed to M. avium but the responses to these antigens were lower than those observed in animals exposed to M. bovis alone. The implication is that responses to M. avium, although providing some immunity, may mask diagnosis of M. bovis infection, even when specific antigens are employed, potentially contributing to disease transmission in the field.
pfmdr1 mutations contribute to quinine resistance and enhance mefloquine and artemisinin sensitivity in Plasmodium falciparum
Authors: Sidhu Amar Bir Singh , Valderramos Stephanie Gaw , Fidock David A.
Published: July 2005

The emergence and spread of multidrug resistant Plasmodium falciparum has severely limited the therapeutic options for the treatment of malaria. With ever‐increasing failure rates associated with chloroquine or sulphadoxine‐pyrimethamine treatment, attention has turned to the few alternatives, which include quinine and mefloquine. Here, we have investigated the role of pfmdr1 3′ coding region point mutations in antimalarial drug susceptibility by allelic exchange in the GC03 and 3BA6 parasite lines. Results with pfmdr1‐recombinant clones indicate a significant role for the N1042D mutation in contributing to resistance to quinine and its diastereomer quinidine. The triple mutations S1034C/N1042D/D1246Y, highly prevalent in South America, were also found to enhance parasite susceptibility to mefloquine, halofantrine and artemisinin. pfmdr1 3′ mutations showed minimal effect on P. falciparum resistance to chloroquine or its metabolite mono‐desethylchloroquine in these parasite lines, in contrast to previously published results obtained with 7G8 parasites. This study supports the hypothesis that pfmdr1 3′ point mutations can significantly affect parasite susceptibility to a wide range of antimalarials in a strain‐specific manner that depends on the parasite genetic background.
Dps/Dpr ferritin‐like protein: insights into the mechanism of iron incorporation and evidence for a central role in cellular iron homeostasis in Streptococcus suis
Authors: Pulliainen Arto T. , Kauko Anni , Haataja Sauli , Papageorgiou Anastassios C. , Finne Jukka
Published: July 2005

The Dps family members constitute a distinct group of multimeric and ferritin‐like iron binding proteins (up to 500 iron atoms/12‐mer) that are widespread in eubacteria and archaea and implicated in oxidative stress resistance and virulence. Despite the wealth of structural knowledge, the mechanism of iron incorporation has remained elusive. Here, we provide evidence on Dpr of the swine and human pathogen Streptococcus suis that: (i) iron incorporation proceeds by Fe(II) binding, Fe(II) oxidation and subsequent storage as Fe(III); (ii) Fe(II) atoms enter the  12‐mer cavity through four hydrophilic pores; and (iii) Fe(II) atoms are oxidized inside the 12‐mer cavity at 12 identical inter‐subunit sites, which are structurally different but functionally equivalent to the ferroxidase centres of classical ferritins. We also provide evidence, by deleting and ectopically overexpressing Dpr, that Dpr affects cellular iron homeostasis. The key residues responsible for iron incorporation in S. suis Dpr are well conserved throughout the Dps family. A model for the iron incorporation mechanism of the Dps/Dpr ferritin‐like protein is proposed.
High‐throughput in planta expression screening identifies a class II ethylene‐responsive element binding factor‐like protein that regulates plant cell death and non‐host resistance
Authors: Nasir Khairun Hisam Bin , Takahashi Yoshihiro , Ito Akiko , Saitoh Hiromasa , Matsumura Hideo , Kanzaki Hiroyuki , Shimizu Takeshi , Ito Minako , Fujisawa Shizuko , Sharma Prakash C. , Ohme‐Takagi Masaru , Kamoun Sophien , Terauchi Ryohei
Published: July 2005

We performed high‐throughput screening using the potato virus X (PVX) system to overexpress Nicotiana benthamiana genes in planta and identify positive regulators of cell death. This screening identified NbCD1, a novel class II ethylene‐responsive element binding factor (ERF), as a potent inducer of the hypersensitive response (HR)‐like cell death. NbCD1 expression was induced by treatments with INF1 elicitor and a non‐host pathogen Pseudomonas cichorii. NbCD1 exhibited transcriptional repressor activity through its EAR motif, and this motif was necessary for NbCD1 to cause cell death. We identified 58 genes that displayed altered transcription following NbCD1 overexpression. NbCD1 overexpression downregulated the expression of HSR203, a negative regulator of hypersensitive death. Conditional expression of NbCD1 in Arabidopsis also caused cell death, indicating that NbCD1 downstream cascades are conserved in dicot plants. To further confirm the role of NbCD1 in defense, we used virus‐induced gene silencing to demonstrate that NbCD1 is required for non‐host resistance of N. benthamiana to the bacterial pathogen P. cichorii. Our data point to a model of transcriptional regulatory cascades. NbCD1 positively regulates cell death and contributes to non‐host resistance, possibly by downregulating the expression of other defense response genes.
Carbon isotope discrimination: potential for screening salinity tolerance in rice at the seedling stage using hydroponics
Authors: Shaheen R. , Hood‐Nowotny R. C.
Published: July 2005

Rice is a moderately salt‐sensitive crop species and soil salinity is the single most widespread soil toxicity problem lacing rice production. The quantification of salinity resistance poses serious problems in the field because of climatic factors and field heterogeneity. In the present study. rice germplasm obtained from the International Rice Research Institute (IRRI). Philippines, was screened in a naturally lit (11 h daylight) glasshouse‐based hydroponics unit at two salinity levels (4 and 6 dS/m), Phenotypic performance based on survival of tolerant, moderately tolerant and susceptible isogenic lines along with tolerant and susceptible parents was evaluated after 10 and 13 days of salt stress. Plants were harvested after second scoring and carbon isotope discrimination in the leaves (A) was measured. δ1 ranged from 19.5 to 22.9%, A highly significant negative correlation (r =−0.95. P < 0.001) between δ and visual scoring was observed. Data indicated the potential of using δ as a physiological indicator for salinity tolerance in rice seedlings grown in hydroponics.
The possibilities of early selection of late‐flowering almonds as a function of seed germination or leafing time of seedlings
Authors: Dicenta F. , García‐Gusano M. , Ortega E. , Martinez‐Gómez P.
Published: July 2005

Obtaining new, late‐flowering almonds by crossing is a very long process, mainly due to the long juvenile period of the seedlings obtained. It would be very useful to have an early selection method to identify the late‐flowering genotypes, which would increase the efficiency of the breeding programmes. In this work, a study was made of the possibility of using the chilling requirements of seeds for germination and the leafing time of seedlings as criteria for early selection of flowering time, in 502 seedlings belonging to 13 families, over a four‐year period. Crossings were carried out in 1997. Seeds were stratified at 7°C. and the number of weeks necessary for germination was determined. Seedlings were planted in 1998. Between 1999 and 2002 the leafing time was recorded, and in 2001 and 2002 the time of flowering was also recorded. The results showed that, although some correlation was observed between the germination or leafing time and the flowering time, these correlations were not strong enough to be used as efficient criteria for early selection of late‐flowering seedlings within each family. for which variability was limited. To obtain late‐flowering cultivars. the best method is to cross progenitors which flower as late as possible, and later select the seedlings according to their flowering time. Leafing time could be used for early selection with certain success when progenitors have very different flowering times (VCTJ early and very late), since the expected variability within families will be very large.
Development of a cleaved amplified polymorphic sequence (CAPS) marker linked to pungency in pepper
Authors: Minamiyama Y. , Kinoshita S. , Inaba K. , Inoue M.
Published: July 2005

The complete tack of pungency in pepper (Capsicum annuum L.) is controlled by a single recessive gene (c). To develop a molecular marker linked to the C locus, two segregating F2 populations (TM2 and TF2) derived from crosses between occasionally pungent and non‐pungent peppers in C. annuum were used. Using the RAPD (random amplified polymorphic DNA) technique in combination with a bulked segregation analysis, two RAPD markers, OPD20‐800 and OPY09‐800, were obtained. Of the two markers, the more closely linked marker. OPY09‐800, was converted into a codominant CAPS (cleaved amplified polymorphic sequence) marker using data from the alignment of the two allelic sequences. This CAPS marker was linked to the C locus (3.6 cM in the TF2 population), and polymorphism was detected among accessions within C. annuum. This marker might be helpful for the selection of a c gene in backcross and progeny tests in a conventional breeding system.