bluuebunny/crossref_metadata_2025 huggingface dataset:
Human T lymphocytes and mast cells differentially express and regulate extra‐ and intracellular CXCR1 and CXCR2
Authors: Lippert Undine , Zachmann Karolin , Henz Beate M. , Neumann Christine
Published: July 2004

Abstract:  CXCL8 plays a major role in cell recruitment to sites of inflammation. Apart from neutrophils, little is known, however, about the cellular distribution and regulation of CXCL8 receptors in cells involved in acquired and adaptive immune responses. In previous studies, we have demonstrated the extracellular expression and function of CXCR1/2 on mast cells and also detected an intracellular pool of CXCR1/2. Here, we have addressed the question of receptor regulation during stimulation of human mast cells (HMC‐1 cell line) and have studied T cells in comparison. Cell permeabilization was performed to detect both surface and possible intracellular receptor pools. HMC‐1 cells stained positive for both receptors on the cell surface (CXCR1, 50%; CXCR2, 51%) and also after cell permeabilization (CXCR1, 86%; CXCR2, 74%). Similarly, T cells exhibited both cell‐surface receptor expression (CXCR1, 30%; CXCR2, 23%) and higher total receptor expression (CXCR1, 50%; CXCR2, 36%), although overall values were lower than that in HMC‐1 cells. On immunoblot, molecular weights of extra‐ and intracellular receptors on mast cells were the same, excluding altered receptor glycosylation. On stimulation with phorbol 12‐myristate 13‐acetate plus calcium ionophore, a time‐dependent decrease of surface‐membrane receptors was observed in both cell types, while total receptor remained the same, suggesting that receptor shedding is not involved. The kinetics of membrane receptor internalization and replenishment differed for the two cell types. Furthermore, receptor internalization was associated with decreased F‐actin polymerization, a basic prerequisite for cell migration. These findings demonstrate for the first time the expression of extra‐ and intracellular CXCR1/2 receptors on T cells and delineate the dynamics of CXCR1/2 receptors on mast cells and T cells. Furthermore, they suggest a cell‐type‐specific and finely tuned regulation of chemokine responses at the receptor level in the context of inflammation.
Characterization of the expression and function of N‐methyl‐D‐aspartate receptor in keratinocytes
Authors: Morhenn V. B. , Murakami M. , O'Grady T. , Nordberg J. , Gallo R. L.
Published: July 2004

Abstract:  The N‐methyl‐D‐aspartate (NMDA) receptor is expressed on neural tissue where it gates calcium ion entry upon stimulation. Using immunohistochemistry, it has been demonstrated in this study that the NMDAR1 receptor is also expressed on keratinocytes (KCs) in normal human skin and inflamed psoriatic skin in vivo. Furthermore, the NMDA receptor was functional as demonstrated by the ability of this receptor to trigger Ca++ influx in KCs. Incubation of cultured, human KCs with MK‐801 decreases the cell growth and induces an increase in apoptosis. These findings demonstrate that the KC expression of NMDA receptor is a mechanism through which the influx of Ca++ into the cell can be regulated and suggest that the expression of this receptor may play a role in the regulation of KC growth and differentiation.
N‐methyl‐D‐aspartate receptors influence the intracellular calcium concentration of keratinocytes
Authors: Fischer Matthias , Glanz Dagobert , William Thomas , Klapperstück Thomas , Wohlrab Johannes , Marsch Wolfgang CH.
Published: July 2004

Abstract: In the present study, the distribution of ionotropic glutamate receptors of the N‐methyl‐D‐aspartate (NMDA)‐receptor type was immunohistochemically demonstrated in healthy human skin (n = 22) and healthy buccal mucosa (n = 20). Moreover, the intracellular calcium concentration of HaCaT‐cells and native human keratinocytes were studied under the influence of the selective agonist NMDA and the selective NMDA‐antagonist MK‐801.

Immunohistochemical imaging of NMDA receptors in healthy epidermis showed a positive reaction in the stratum basale, spinosum and granulosum, whereby the greatest expression was observed in the granular layer. In the mucosal preparations, the distribution of NMDA receptors was observed to be equal in all cell layers. In the cell culture (HaCaT‐cells), NMDA concentrations between 25 µM and 1 mM resulted in a significant increase in the number of cells showing elevated intracellular calcium concentration. This effect could be significantly reduced by prior application of MK‐801 (100 µM). In supplementary tests on HaCaT‐keratinocytes, blockade of the keratinocytic NMDA receptors with MK‐801 suppressed the differentiation of the cells (expression of cytokeratin 10). The proliferation of cells was not influenced by NMDA.

The investigations showed that glutamate receptors of the NMDA type have an influence on keratinocytic calcium concentration. This appears especially important for the differentiation of keratinocytes.
Pathological and immunological profiles of rat tuberculosis
Authors: Sugawara Isamu , Yamada Hiroyuki , Mizuno Satoru
Published: July 2004

To investigate the pathological and immunological profiles of rat tuberculosis, Lewis female rats were infected aerially with Mycobacterium tuberculosis. Histopathology, immunological profiles of mononuclear cells from M. tuberculosis‐infected rat lung tissue, and the expression patterns of cytokine and iNOS mRNAs were examined over time. M. tuberculosis induced granulomatous lesions in the lungs, spleen, lymph nodes and liver, but these lesions lacked central necrosis. Multinucleate giant cells were observed in late‐phase tuberculosis. CD4+ and CD8+ T cells increased with time and reached a peak 5 weeks after infection, decreasing gradually thereafter. ED1 antigen, suggestive of alveolar macrophages, was expressed at a high level in early phase tuberculosis and remained at the same level even in the late phase. OX62 antigen increased gradually and reached a peak 5 weeks after infection. Interferon‐γ, tumour necrosis factor‐α and iNOS mRNAs were expressed strongly over time, but their expression decreased 12 weeks after infection. Because rat tuberculosis is very similar to murine tuberculosis and it is easy to obtain mononuclear cells from M. tuberculosis‐infected rat lung tissue, the rat tuberculosis model appears to be suitable for immunological studies in vivo.
Adult stem cell plasticity: will engineered tissues be rejected?
Authors: Fang Te‐Chao , Alison Malcolm R. , Wright Nicholas A. , Poulsom Richard
Published: July 2004

The dogma that adult tissue‐specific stem cells remain committed to supporting only their own tissue has been challenged; a new hypothesis, that adult stem cells demonstrate plasticity in their repertoires, is being tested. This is important because it seems possible that haematopoietic stem cells, for example, could be exploited to generate and perhaps deliver cell‐based therapies deep within existing nonhaematopoietic organs.

Much of the evidence for plasticity derives from histological studies of tissues from patients or animals that have received grafts of cells or whole organs, from a donor bearing (or lacking) a definitive marker. Detection in the recipient of appropriately differentiated cells bearing the donor marker is indicative of a switch in phenotype of a stem cell or a member of a transit amplifying population or of a differentiated cell. In this review, we discuss evidence for these changes occurring but do not consider the molecular basis of cell commitment.

In general, the extent of engraftment is low but may be increased if tissues are damaged. In model systems of liver regeneration, the repeated application of a selection pressure increases levels of engraftment considerably; how this occurs is unclear. Cell fusion plays a part in regeneration and remodelling of the liver, skeletal muscle and even regions of the brain.

Genetic disease may be amenable to some forms of cell therapy, yet immune rejection will present challenges. Graft‐vs.‐host disease will continue to present problems, although this may be avoided if the cells were derived from the recipient or they were tolerized. Despite great expectations for cellular therapies, there are indications that attempts to replace missing proteins could be confounded simply by the development of specific immunity that rejects the new phenotype.
In situ analysis of lung antigen‐presenting cells during murine pulmonary infection with virulent Mycobacterium tuberculosis
Authors: Pedroza‐González Alexander , García‐Romo Gina S. , Aguilar‐León Diana , Calderon‐Amador Juana , Hurtado‐Ortiz Raquel , Orozco‐Estevez Hector , Lambrecht Bart N. , Estrada‐García Iris , Hernández‐Pando Rogelio , Flores‐Romo Leopoldo
Published: July 2004

Scarce information exists about the role of lung antigen‐presenting cells (APCs) in vivo during pulmonary tuberculosis. As APCs activate cellular immunity, following intratracheal inoculation with virulent Mycobacterium tuberculosis, we assessed in situ lung APC recruitment, distribution, granuloma involvement, morphology and mycobacterial burden by using MHC‐CII, CD14, scavenger receptor class A (SRA), the murine dendritic cell (DC)‐restricted marker CD11c and Ziehl–Neelsen staining. CD11c+ DC and CD14+ cell recruitment into lungs appeared by day 14, continuing until day 60. MHC‐CII+ cells increased since day 7, persisting until day 60. Thus, virulent mycobacteria delays (14–21 days) lung APC recruitment compared to model antigens and nonvirulent bacilli (24–48 h).

Regarding granuloma constitution, highly bacillary CD14+ and SRA+ cells were centrally located. MHC‐CII+ cells were more peripheral, with less mycobacteria. CD11c+ cells were heterogeneously distributed within granulomas, with scarce bacilli.

When labelling lung suspensions for MHC‐CII and classifying cells as macrophages or DC, then staining for Ziehl–Neelsen, a remarkable segregation was found regarding bacillary burden. Most macrophage‐like cells contained numerous bacilli, while DC had no or scarce mycobacteria. This implies differential APC contributions in situ during pulmonary tuberculosis regarding mycobacterial uptake, granuloma involvement and perhaps bacillary growth.
Seroconversion and Seroreversion of Helicobacter pylori Antibodies Over a 9‐Year Period and Related Factors in Japanese Adults
Authors: Kikuchi Shogo , Ohgihara Atsushi , Hasegawa Ayako , Miki Kazumasa , Kaneko Eizo , Mizukoshi Hiroshi
Published: July 2004

Background.  There are still insufficient data on the frequency of seroconversion and seroreversion of Helicobacter pylori antibodies. The frequency of serochange and related factors were investigated in this study over 9 years.

Subjects and methods.  Using sera from 3104 workers who underwent health checks in 1989 and again in 1998, H. pylori antibodies were measured. Those with intermediate serology were excluded from the study. Information on past history was collected using a questionnaire.

Results.  Of the 912 seronegative and 1286 seropositive subjects in 1989, seroconversion was observed in 57 and seroreversion in 91 subjects. Seroconversion and seroreversion rates over the 9‐year period were 6.3% and 7.1%, respectively, giving rates per 1000 person‐years (with 95% confidence interval) of 7.0 (5.2–8.7) and 7.9 (6.3–9.4), respectively. Subjects that reported abdominal symptoms or gastric fiberscope use showed significantly higher seroconversion rates than controls (8.7 vs. 4.5 and 9.2 vs. 1.6, respectively), which remained significant after adjustment for age and gender. Those with a history of duodenal ulcers, a smoking habit or a drinking habit showed significantly lower seroreversion rates than controls (3.5 vs. 8.9, 5.4 vs. 9.2 and 5.9 vs. 13.3, respectively). After adjustment, the association between seroreversion and smoking habit disappeared, while the associations with history of duodenal ulcers and drinking habit remained.

Conclusions.  Those with a history of nonspecific abdominal symptoms and those with a history of gastric fiberscope use showed higher seroconversion rates. Alcohol consumption and duodenal ulcers may inhibit the autoeradication of H. pylori, possibly through increased acid secretion.
IgG Subclass Responses in Childhood Helicobacter pylori Duodenal Ulcer: Evidence of T‐Helper Cell Type 2 Responses
Authors: Campbell David I. , Pearce Mark S. , Parker Louise , Thomas Julian E.
Published: July 2004

Background.  Duodenal ulcer in adults chronically infected with Helicobacter pylori is associated with a polarized T‐helper cell type 1 (Th1) mucosal immune response, with a predominantly immunoglobulin G2 (IgG2) systemic specific response. It has been suggested that children colonized by H. pylori also produce a mucosal Th1 response, but there are few studies that have measured IgG subclass responses in children with duodenal ulcer.

Materials and methods.  Seven children with endoscopically proven duodenal ulcer and H. pylori infection and 18 children with biopsy proven H. pylori infection but no duodenal ulcer had relative concentrations of IgG subclass responses (IgGsc) against H. pylori antigens measured by ELISA. Eighteen IgG seropositive adults acted as controls. The range of antigens recognised by IgG1 and IgG2 subclass responses were investigated by Western blots.

Results.  There were no differences in mean IgGsc responses between children with or without duodenal ulcer. Adults produced an IgG2 predominant response. Western blots showed no qualitative differences in antigens recognised by IgG1 or IgG2.

Conclusion.  Children with duodenal ulcer, in contrast to adults, produce an IgGsc response consistent with a mucosal Th2 response to H. pylori regardless of the presence of duodenal ulceration. This suggests that disease causation amongst children with H. pylori associated duodenal ulceration may not be dependant upon a mucosal Th1 biased response.
Stool Antigen Test for the Diagnosis of Helicobacter pylori Infection: a Systematic Review
Authors: Gisbert Javier P. , Pajares José María
Published: July 2004

Our aim was to review systematically the diagnostic accuracy of the Helicobacter pylori stool antigen test. Bibliographical searches were performed in several electronic databases and abstracts from congresses up to May 2003. Eighty‐nine studies (10,858 patients) evaluated the stool antigen test in untreated patients. Mean sensitivity, specificity, positive predictive value and negative predictive value were 91%, 93%, 92% and 87%, respectively. Analysis of the eight studies (1399 patients) in which pretreatment evaluation of the monoclonal stool antigen test was performed showed better (p < .001) results (96%, 97%, 96% and 97%, respectively), with a clearer distinction between positive and negative results. Thirty‐nine studies (3147 patients) evaluated the stool antigen test for the confirmation of H. pylori eradication 4–8 weeks after therapy, with accuracies of 86%, 92%, 76% and 93% for mean sensitivity, specificity, positive predictive value and negative predictive value, respectively. Results were similar when a gold standard based on at least two methods was used. Relatively low accuracy was reported in some posttreatment studies with the polyclonal stool antigen test. However, excellent results (p < .001) were achieved in all the six studies evaluating the monoclonal stool antigen test 4–8 weeks posttreatment. Results evaluating the stool antigen test < 4 weeks posttreatment are contradictory. Proton‐pump inhibitors seem to affect the accuracy of the stool antigen test. Sensitivity and/or specificity in patients with gastrointestinal bleeding may be suboptimal. The stool antigen test performs well in children. Finally, the stool antigen test seems to be a cost‐effective method.
Comparative Chemical and Biological Characterization of the Lipopolysaccharides of Gastric and Enterohepatic Helicobacters
Authors: Hynes Sean O. , Ferris John A. , Szponar Bogumila , Wadström Torkel , Fox James G. , O'Rourke Jani , Larsson Lennart , Yaquian Elisa , Ljungh Åsa , Clyne Marguerite , Andersen Leif P. , Moran Anthony P.
Published: July 2004

Background.  The lipopolysaccharide of Helicobacter pylori plays an important role in colonization and pathogenicity. The present study sought to compare structural and biological features of lipopolysaccharides from gastric and enterohepatic Helicobacter spp. not previously characterized.

Materials and methods.  Purified lipopolysaccharides from four gastric Helicobacter spp. (H. pylori, Helicobacter felis, Helicobacter bizzozeronii and Helicobacter mustelae) and four enterohepatic Helicobacter spp. (Helicobacter hepaticus, Helicobacter bilis, ‘Helicobacter sp. flexispira’ and Helicobacter pullorum) were structurally characterized using electrophoretic, serological and chemical methods.

Results.  Structural insights into all three moieties of the lipopolysaccharides, i.e. lipid A, core and O‐polysaccharide chains, were gained. All species expressed lipopolysaccharides bearing an O‐polysaccharide chain, but H. mustelae and H. hepaticus produced truncated semirough lipopolysaccharides. However, in contrast to lipopolysaccharides of H. pylori and H. mustelae, no blood group mimicry was detected in the other Helicobacter spp. examined. Intra‐species, but not interspecies, fatty acid profiles of lipopolysaccharides were identical within the genus. Although shared lipopolysaccharide‐core epitopes with H. pylori occurred, differing structural characteristics were noted in this lipopolysaccharide region of some Helicobacter spp. The lipopolysaccharides of the gastric helicobacters, H. bizzozeronii and H. mustelae, had relative Limulus amoebocyte lysate activities which clustered around that of H. pylori lipopolysaccharide, whereas H. bilis, ‘Helicobacter sp. flexispira’ and H. hepaticus formed a cluster with approximately 1000–10,000‐fold lower activities. H. pullorum lipopolysaccharide had the highest relative Limulus amoebocyte lysate activity of all the helicobacter lipopolysaccharides (10‐fold higher than that of H. pylori lipopolysaccharide), and all the lipopolysaccharides of enterohepatic Helicobacter spp. were capable of inducing nuclear factor‐Kappa B(NF‐κB) activation.

Conclusions.  The collective results demonstrate the structural heterogeneity and pathogenic potential of lipopolysaccharides of the Helicobacter genus as a group and these differences in lipopolysaccharides may be indicative of adaptation of the bacteria to different ecological niches.
Helicobacter pylori Infection and Immune Thrombocytopenic Purpura: an Update
Authors: Franchini Massimo , Veneri Dino
Published: July 2004

Data are accumulating on the association between Helicobacter pylori infection and idiopathic thrombocytopenic purpura (ITP) and the significant increase in platelet count after bacterial eradication. The aim of this review was to consider the studies so far published on H. pylori infection and ITP in order to evaluate a possible correlation between these two conditions. A review of the literature showed that 278 out of the 482 ITP patients investigated (58%) were positive for H. pylori infection and that the bacterium was eradicated in 88% of cases. Eradication therapy was accompanied by a complete or partial platelet response in approximately half the cases. Overall, these data show that H. pylori eradication in patients with ITP is effective in increasing platelet count. However, because the studies so far published are few, are sometimes controversial and involve small series of patients, further studies on larger numbers of patients with longer follow‐up are needed to confirm these preliminary findings.
Polymorphism of the Helicobacter pylori feoB Gene in Korea: a Possible Relation with Iron‐Deficiency Anemia?
Authors: Jeon Byung Ha , Oh Yoo Joung , Lee Na Gyong , Choe Yon Ho
Published: July 2004

Background.  Helicobacter pylori is a causative agent of gastritis, and H. pylori infection is thought to be correlated with iron‐deficiency anemia (IDA) at puberty. The H. pylori feoB gene product, a high‐affinity ferrous iron transporter, plays a central role in iron acquisition and virulence. This study was undertaken to analyze H. pylori feoB status according to clinical data, including antral gastritis with or without IDA.

Methods.  Fourteen H. pylori‐positive patients aged from 10 to 18 years were categorized into subgroups based on the presence or absence of IDA. Eight patients were diagnosed as having IDA; the other six showed normal hematological findings. Genomic DNA was isolated from H. pylori cultured from each gastric biopsy specimen. Five sets of primers were used for the PCR amplification of the feoB gene. Linking and sequencing of PCR products generated the feoB region, which was 1.93 kb in size. The feoB gene sequences of H. pylori J99 and 26695 were compared with the clinical strains, and the sequences of feoB regions in the IDA (+) and (–) groups were compared.

Results.  Sequence analysis of the complete coding region of the feoB gene revealed 16 sites of polymorphism or mutation. Among these, three polymorphisms (E/T254A, I263V, and K511Q) were indigenous to the Korean clinical strains. Although statistically significant differences were observed at four sites (K127T, A273S/P, I438V and I441T) between IDA (+) and (–), the number of specimens was too low to assess the significance of the differences.

Conclusion.  The four polymorphisms of the feoB gene observed appear to be related to the clinical phenotype of IDA, but the relation is unclear because of the small number of strains studied. Further studies are required to confirm a correlation between IDA and H. pylori infection.
Dual vs. Triple Therapy for Childhood Helicobacter pylori Gastritis: a Double‐Blind Randomized Multicentre Trial
Authors: Oderda Giuseppina , Marinello Daniela , Lerro Pietro , Kuvidi Maria , De’Angelis Gian Luigi , Ferzetti Anna , Cucchiara Salvatore , Franco Maria Teresa , Romano Claudio , Strisciuglio Pietro , Pensabene Lucia
Published: July 2004

Background.  Data on the efficacy of eradication treatment for Helicobacter pylori gastritis in children are scarce.

Aim.  To evaluate the efficacy of triple therapy with lansoprazole plus amoxicillin and tinidazole vs. dual therapy with amoxicillin and tinidazole in a double‐blind randomized multicentre trial, and the usefulness of eradication in terms of long‐term symptom resolution.

Subjects.  We enrolled 43 consecutive children undergoing endoscopy for upper gastrointestinal dyspepsia with H. pylori gastritis. They underwent a 13C‐urea breath test, completed a 2‐week symptom diary card, and were randomized. Treatment was given in a Redidose box (Redidose Company Ltd., Brighton, UK) containing either lansoprazole‐amoxicillin‐tinidazole (triple therapy) or placebo plus amoxicillin‐tinidazole (dual therapy) for 1 week. The completion of a 2‐week symptom diary card and the performance of a breath test were repeated 6 weeks and 6 months after the end of therapy. One to two years later, a structured telephone interview was conducted with 36 of the children.

Results.  According to the breath test, 6 weeks after the end of therapy H. pylori was eradicated in 15 of 22 children on triple therapy [68.2%; 95% confidence interval (CI) = 45–88] and in 15 of 21 children on dual therapy (71%; 95% CI = 48–89; not significant), and 6 months after the end of therapy it was eradicated in 16 of 22 children on triple therapy (72.7%) and in 15 of 21 children on dual therapy. Six months after therapy, symptoms were analysed in 11 H. pylori‐positive and 31 H. pylori‐negative children, and it was found that dyspeptic symptoms had disappeared or improved in both groups, with no difference between them. One to two years later, 36 children were interviewed. Epigastric pain had recurred in three of 26 H. pylori‐negative and in seven of 10 H. pylori‐positive children (p = .001); in three of the latter, pain was severe and required additional treatment.

Conclusion.  One‐week triple or dual therapy with two antibiotics achieved similar eradication rates. Soon after treatment, symptoms disappeared or improved in most children irrespective of eradication, but epigastric pain recurred in the majority of the still‐infected children within 2 years.
Interaction of Rho‐kinase with myosin II at stress fibres
Authors: Kawabata Saeko , Usukura Jiro , Morone Nobuhiro , Ito Masaaki , Iwamatsu Akihiro , Kaibuchi Kozo , Amano Mutsuki
Published: July 2004

Rho‐kinase and myosin phosphatase cooperatively regulate the phosphorylation level of myosin light chain and are involved in the formation of stress fibres and smooth muscle contraction. Rho‐kinase has been known to be localized at stress fibres, but little is known about the mechanism of its localization. Here we identified non‐muscle myosin heavy chain IIA and IIB as the pleckstrin homology domain‐interacting molecules by affinity column chromatography. The pleckstrin homology domain of Rho‐kinase binds to myosin II directly in in vitro cosedimentation assay. The C‐terminal region of the pleckstrin homology domain was important for this interaction, and the point mutations in the pleckstrin homology domain mutant (W1170A, W1340L) resulted in a decrease in the binding. We also found that the pleckstrin homology domain, but not the pleckstrin homology domain mutant (W1170A, W1340L), was localized at stress fibres in fibroblasts. These results indicate that Rho‐kinase is localized at stress fibres through binding of the pleckstrin homology domain to myosin II.
The crystal structure of microtubule‐associated protein light chain 3, a mammalian homologue of Saccharomyces cerevisiae Atg8
Authors: Sugawara Kenji , Suzuki Nobuo N. , Fujioka Yuko , Mizushima Noboru , Ohsumi Yoshinori , Inagaki Fuyuhiko
Published: July 2004

Microtubule‐associated protein light chain 3 (LC3), a mammalian homologue of yeast Atg8, plays an essential role in autophagy, which is involved in the bulk degradation of cytoplasmic components by the lysosomal system. Here, we report the crystal structure of LC3 at 2.05 Å resolution with an R‐factor of 21.8% and a free R‐factor of 24.9%. The structure of LC3, which is similar to those of Golgi‐associated ATPase enhancer of 16 kDa (GATE‐16) and GABAA receptor‐associated protein (GABARAP), contains a ubiquitin core with two α helices, α1 and α2, attached at its N‐terminus. Some common and distinct features are observed among these proteins, including the conservation of residues required to form an interaction among α1, α2 and the ubiquitin core. However, the electrostatic potential surfaces of these helices differ, implicating particular roles to select specific binding partners. Hydrophobic patches on the ubiquitin core of LC3, GABARAP and GATE‐16 are well conserved and are similar to the E1 binding surface of ubiquitin and NEDD8. Therefore, we propose that the hydrophobic patch is a binding surface for mammalian Atg7 similar to a ubiquitin‐like conjugation system. We also propose the functional implications of the ubiquitin fold as a recognition module of target proteins.
Fluctuation of chromatin unfolding associated with variation in the level of gene expression
Authors: Sato Noriko , Nakayama Masahito , Arai Ken‐ichi
Published: July 2004

We examined whether spontaneous alteration of chromatin structure, if any, correlates with variation in gene expression. Gene activation is associated with changes in chromatin structure at different levels. Large‐scale chromatin unfolding is one such change detectable under the light microscope. We established cell clones carrying tandem repeats (more than 50 copies spanning several hundred kb) of the GFP (green fluorescent protein)‐ASK reporter genes driven by a tetracycline responsive promoter. These clones constitutively express the transcriptional transactivator. Flow cytometry and live‐recording fluorescence microscopy revealed that, although fully activated by a saturating amount of doxycycline, GFP‐ASK expression fluctuated in individual cell clones, regardless of the cell cycle stage. The GFP‐ASK expression changed from lower to higher levels and vice versa within a few cell cycles. Furthermore, the levels of GFP‐ASK expression were correlated with the degrees of chromatin unfolding of the integrated array as detected by FISH (fluorescent in situ hybridization). The chromatin unfolding was not coupled to a mitotic event; around one‐third of the daughter‐pairs exhibited dissimilar degrees of chromatin unfolding. We concluded that fluctuation of chromatin unfolding was likely to result in variation in gene expression, although the source of the fluctuation of chromatin unfolding remains to be studied.
Nrf2 deficiency causes tooth decolourization due to iron transport disorder in enamel organ
Authors: Yanagawa Toru , Itoh Ken , Uwayama Junya , Shibata Yasuaki , Yamaguchi Akira , Sano Tsuneyoshi , Ishii Tetsuro , Yoshida Hiroshi , Yamamoto Masayuki
Published: July 2004

Rodents have brownish‐yellow incisors whose colour represents their iron content. Iron is deposited into the mature enamel by ameloblasts that outline enamel surface of the teeth. Nrf2 is a basic region‐leucine zipper type transcription factor that regulates expression of a range of cytoprotective genes in response to oxidative and xenobiotic stresses. We found that genetically engineered Nrf2‐deficient mice show decolourization of the incisors. While incisors of wild‐type mice were brownish yellow, incisors of Nrf2‐deficient mice were greyish white in colour. Micro X‐ray imaging analysis revealed that the iron content in Nrf2‐deficient mouse incisors were significantly decreased compared to that of wild‐type mice. We found that iron was aberrantly deposited in the papillary layer cells of enamel organ in Nrf2‐deficient mouse, suggesting that the iron transport from blood vessels to ameloblasts was disturbed. We also found that ameloblasts of Nrf2‐null mouse show degenerative atrophy at the late maturation stage, which gives rise to the loss of iron deposition to the surface of mature enamel. Our results thus demonstrate that the enamel organ of Nrf2‐deficient mouse has a reduced iron transport capacity, which results in both the enamel cell degeneration and disturbance of iron deposition on to the enamel surface.
Transcriptional responses to epigallocatechin‐3 gallate in HT 29 colon carcinoma spheroids
Authors: McLoughlin Patricia , Roengvoraphoj Monic , Gissel Cornelia , Hescheler Jürgen , Certa Ulrich , Sachinidis Agapios
Published: July 2004

Catechins have been reported to possess anti‐cancer activity in vitro and in vivo. To identify target genes that may be involved in the anti‐tumorigenic effect of catechins, gene expression profiles in adherent human HT 29 colon carcinoma cells, in HT 29 spheroids and in epigallocatechin‐3 gallate (EGCG)‐treated HT 29 cells have been analysed by high‐density oligonucleotide microarrays. Treatment of HT 29 cells with EGCG (2.5–50 µm) resulted in a dose‐dependent inhibition of spheroid formation of HT 29 cells. Forty transcripts were induced at least twofold in 3‐day‐old spheroids relative to normal adherent cells using three replicates. Oncogenes like c‐fos and c‐jun are significantly up‐regulated in spheroids. We identified several signal transduction and proliferation genes which are down‐regulated in response to EGCG treatment. Increase in the mRNA expression profile of c‐Fos correlated well with protein levels in HT 29 spheroids whereas EGCG did not affect protein formation. In agreement with the DNA chip data, IQGAP2 protein was not increased in spheroids but protein formation was totally blocked in EGCG‐treated cells. Interestingly, no change in expression of cytotoxic or apoptotic related genes has been observed in EGCG‐treated cells. Our findings suggest that EGCG may exert its anti‐cancer activity through modulation of expression of a number of genes that are involved in cell proliferation, cell‐cell contacts and cell‐matrix interactions.
Do smokers know how to quit? Knowledge and perceived effectiveness of cessation assistance as predictors of cessation behaviour
Authors: Hammond D. , McDonald P. W. , Fong G. T. , Borland R.
Published: July 2004

Aims  Despite the existence of effective cessation methods, the vast majority of smokers attempt to quit on their own. To date, there is little evidence to explain the low adoption rates for effective forms of cessation assistance, including pharmaceutical aids. This study sought to assess smokers’ awareness and perceived effectiveness of cessation methods and to examine the relationship of this knowledge to cessation behaviour.

Design  A random‐digit‐dial telephone survey (response rate = 76%) with 3‐month follow‐up was conducted with 616 adult daily smokers in South‐Western Ontario, Canada.

Measurements  A baseline survey assessed smoking behaviour, as well as smokers’ awareness and perceived effectiveness of cessation assistance. A follow‐up survey measured changes in smoking behaviour and adoption of cessation assistance at 3 months.

Findings  Participants demonstrated a poor recall of cessation methods: 45% of participants did not recall nicotine gum, 33% did not recall the nicotine patch and 57% did not recall bupropion. Also, many participants did not believe that the following cessation methods would increase their likelihood of quitting: nicotine replacement therapies (36%), bupropion (35%), counselling from a health professional (66%) and group counselling/quit programmes (50%). In addition, 78% of smokers indicated that they were just as likely to quit on their own as they were with assistance. Most important, participants who perceived cessation methods to be effective at baseline, were more likely to intend to quit (OR = 1.80, 95% CI: 1.12–2.90), make a quit attempt at follow‐up (OR = 1.80, 95% CI: 1.03–3.16) and to adopt cessation assistance when doing so (OR = 3.62, 95% CI: 1.04–12.58).

Conclusions  This research suggests that many smokers may be unaware of effective cessation methods and most underestimate their benefit. Further, this lack of knowledge may represent a significant barrier to treatment adoption.
Prevalence of non‐medical drug use and dependence among homosexually active men and women in the US population
Authors: Cochran Susan D. , Ackerman Deborah , Mays Vickie M. , Ross Michael W.
Published: July 2004

Aims The aim of this study was to compare patterns of drug use and dependence between homosexually experienced and exclusively heterosexually experienced individuals.

Design We used a cross‐sectional national household interview survey conducted in the United States.

Setting Secondary data analysis of the 1996 National Household Survey on Drug Abuse was employed.

Participants Participants were sexually active individuals, aged 18 years and older, who reported the genders of their sexual partners in the past 12 months; included 174 homosexually experienced (98 men, 96 women) and 9714 exclusively heterosexually experienced (3922 men, 5792 women) respondents.

Measurements Life‐time, past 30 days and daily use of nine classes of drugs. Symptoms of dysfunctional use and dependence.

Findings There were consistent patterns of elevated drug use in homosexually experienced individuals for life‐time drug use, but these were greatly attenuated for recent use. Homosexually experienced men were more likely to report use of marijuana, cocaine and heroin, and homosexually experienced women more likely to report use of marijuana and analgesics than individuals reporting only opposite‐sex partners. Both homosexually active men and women were more likely than exclusively heterosexually active respondents to report at least one symptom indicating dysfunctional drug use across all drug classes, and to meet criteria for marijuana dependence syndrome. The only difference between homosexually experienced men and women was that men were more likely to report any daily drug use.

Conclusions These data are consistent with surveys suggesting that there is a moderate elevation of drug, particularly marijuana, use and dependence in gay and bisexual men and women when compared to heterosexual men and women.
The effect of matching comprehensive services to patients’ needs on drug use improvement in addiction treatment
Authors: Friedmann Peter D. , Hendrickson James C. , Gerstein Dean R. , Zhang Zhiwei
Published: July 2004

Aims  To examine whether need–service matching in addiction treatment leads to improvements in drug use, and whether treatment duration mediates those improvements.

Design, participants, measurements  This analysis utilizes prospective data from a US cohort of addiction treatment patients who reported service needs beyond core rehabilitative services (n = 3103). ‘Drug use improvement’ is the difference between the patient's peak drug use frequency (in days per month) in the year before intake and in the year after treatment. Overall and primary use of the major illicit drugs (heroin, powder or crack cocaine and marijuana) are considered separately. ‘Need–service match’ means that a patient rated a service as important at intake and reported its receipt during treatment. ‘Percentage of needs matched’ indicates the proportion of five service domains (medical, mental health, family, vocational and housing) so matched.

Findings  In mixed regression models controlling for multiple factors, a greater percentage of needs matched tended to improve primary (β = 0.028, P = 0.09) and overall (β = 0.049, P = 0.05) drug use in the follow‐up year. Exclusion of treatment duration as a covariate doubled the magnitude of these coefficients. The benefits of matching were concentrated among the half of patients reporting needs in four to five rather than one to three domains, and were strongest among patients in long‐term residential facilities. Addressing vocational and housing needs exerted the greatest effects.

Conclusions  Matching comprehensive services to needs is a useful addiction treatment practice, especially for high‐need patients. Treatment duration might partially mediate its effect.
Mortality risk among new onset injection drug users
Authors: Vlahov David , Wang Cun‐lin , Galai Noya , Bareta Joseph , Mehta Shruti H. , Strathdee Steffanie A. , Nelson Kenrad E.
Published: July 2004

Aims  To characterize mortality experience among those who only recently started injection.

Design  Prospective study.

Setting  Independent study clinic within high drug use neighborhoods.

Participants  In 1988–1989, we enrolled 256 adult injection drug users (IDUs) recruited through street outreach who had initiated injection within the prior 2 years.

Measurements  Consenting participants underwent venipuncture for HIV antibody testing and interviews. We prospectively ascertained date and cause of death through follow‐up contact and registry linkages. Analyses included standardized mortality ratios (SMRs) with local, state and national mortality data, adjusted for age, gender and race.

Findings  Baseline median age was 30 years, 70% were male, 95% were African‐American and 90% injected within the prior 6 months. We identified 69 deaths through October 2000; mortality rate was 3.3/100 person‐years. The adjusted SMR with the USA (and Baltimore) as the reference for IDUs was 4.40 (2.43) for 1991–1992, which increased to 8.12 (4.13) by 1993–1994, decreased to 4.43 (2.13) by 1997–1998 and increased slightly to 5.35 (2.79) during 1999–2000. Excluding HIV‐related mortality, SMRs remained elevated. Decline in SMRs was not linked to drug abuse treatment.

Conclusions  These data demonstrate excess mortality among new‐onset IDUs compared with demographically similar peers in the general population, indicating the need for interventions to prevent premature death among young IDUs.
A critical review of the causes of death among post‐mortem toxicological investigations: analysis of 34 buprenorphine‐associated and 35 methadone‐associated deaths
Authors: Pirnay S. , Borron S. W. , Giudicelli C. P. , Tourneau J. , Baud F. J. , Ricordel I.
Published: July 2004

Aims  To assess the trends in the number, mortality and the nature of forensic cases involving toxicological detection of buprenorphine or methadone among toxicological investigations performed in Paris from June 1997 to June 2002.

Design  Retrospective, 5 year study with review of premortem data, autopsy, police reports, hospital data, and post‐mortem toxicological analyses.

Setting and participants  34 forensic cases of buprenorphine and 35 forensic cases of methadone detection among 1600 toxicological investigations performed at the Laboratory of Toxicology in the Medical Examiner's Office in Paris.

Measurements and results  Therapeutic, toxic or lethal drug concentrations were defined based upon the results of blood analyses and the published literature. Drug concentrations were cross‐referenced with other available ante‐ and post‐mortem data. Subsequently, we classified a ‘clear responsibility’, ‘possible responsibility’ or ‘not causative’ role for buprenorphine or methadone in the death process, or ‘no explanation of death’. Buprenorphine and methadone can be regarded as being directly implicated in, respectively, four of 34 death cases (12%) and three of 35 death cases (9%), and their participation in the lethal process is strongly plausible in eight (buprenorphine) and 11 (methadone) additional deaths.

Conclusions  Analysis of causes of death reveals the difficulties in determining the role of substitution drugs in the death process, as many other factors may be involved, including circumstances surrounding death, past history, differential selection of subjects

into either substitution modality and concomitant intake of other drugs (especially benzodiazepines and neuroleptics). The potential for synergistic or additive actions by other isolated molecules—particularly opioids, benzodiazepines, other psychotropes and alcohol—must be also considered.
Under‐reporting of alcohol consumption in household surveys: a comparison of quantity–frequency, graduated–frequency and recent recall
Authors: Stockwell Tim , Donath Susan , Cooper‐Stanbury Mark , Chikritzhs Tanya , Catalano Paul , Mateo Cid
Published: July 2004

Aim  To compare alternative survey methods for estimating (a) levels of at risk alcohol consumption and (b) total volume of alcohol consumed per capita in comparison with estimates from sales data and to investigate reasons for under‐reporting.

Setting  The homes of respondents who were eligible and willing to participate.

Participants  A total of 21 674 Australians aged 14 years and older.

Design  A 2001 national household survey of drug use, experiences and attitudes with weights applied for age, sex, geographic location and day of week of interview.

Measures  Self‐completion questionnaire using quantity–frequency (QF) and graduated–frequency (GF) methods plus two questions about consumption ‘yesterday’: one in standard drinks, another with empirically based estimates of drink size and strength.

Results  The highest estimate of age 14 + per capita consumption of 7.00 l of alcohol derived from recall of consumption ‘yesterday’ or 76.8% of the official estimate. The lowest was QF with 49.8%. When amount consumed ‘yesterday’ was recalled in standard drinks this estimate was 5.27 l. GF questions yielded higher estimates than did QF questions both for total volume (5.25 versus 4.54 l) and also for the proportion of the population at risk of long‐term alcohol‐related harm (10.6%versus 8.1%). With the detailed ‘yesterday’ method 61% of all consumption was on high risk drinking days.

Conclusions  Questions about typical quantities of alcohol consumed can lead to underestimates, as do questions about drinking ‘standard drinks’ of alcohol. Recent recall methods encourage fuller reporting of volumes plus more accurate estimates of unrecorded consumption and the proportion of total alcohol consumption that places drinkers at risk of harm. However, they do not capture longer‐term drinking patterns. It is recommended that both recent recall and measures of longer‐term drinking patterns are included in national surveys.
An expanded comparative map of bovine chromosome 27 targeting dairy form QTL regions*
Authors: Connor E. E. , Sonstegard T. S. , Ashwell M. S. , Bennett G. L. , Williams J. L.
Published: July 2004

At present, the density of genes on the bovine maps is extremely limited and current resolution of the human–bovine comparative map is insufficient for selection of candidate genes controlling many economic traits of interest in dairy cattle. This study describes the chromosomal mapping of 10 selected gene‐associated markers to bovine linkage and radiation hybrid maps to improve the breakpoint resolution in the human–bovine comparative map near two previously identified quantitative trait loci for the linear type trait, dairy form. Two regions of conserved synteny not previously described are reported between the telomeric region of bovine chromosome 27 (BTA27) and human chromosome 3 (HSA3) p24 region and between the HSA4q34.1 region and BTA8. These data increase the number of genes positioned on the bovine gene maps, refine the human–bovine comparative map, and should improve the efficiency of candidate gene selection for the dairy form trait in cattle.
Impact of the ESR gene on litter size and production traits in Czech Large White pigs
Authors: Goliášová E. , Wolf J.
Published: July 2004

To evaluate the effect of the PvuII polymorphism of the oestrogen receptor gene on litter size and production traits in Czech Large White swine, data from 1250 sows and 3600 litters were analysed with two four‐trait animal models. The traits in the first model were number of piglets born alive in a sow's first litter, number of piglets born alive in second and subsequent litters, lifetime daily gain and lean meat percentage. The second model included number of piglets born, number of piglets born alive, number of piglets weaned and litter weight at weaning from first and subsequent litters. The oestrogen receptor (ESR) locus significantly affected prolicacy in the first parity and averaged over all parities (P < 0.05), with allele A superior to allele B. In the first parity, AA sows produced approximately 0.5 more live piglets per litter than BB sows. Averaged over all parities, this difference was c. 0.25 piglets. Results for total number of piglets born and number of piglets weaned were similar to results for numbers born alive. No significant dominance effect was found for prolificacy traits. For litter weight at weaning, no significant additive effect was observed at the ESR locus, but a significant negative dominance effect (−1.5 kg) was estimated averaged across parities (litters of AB sows were similar to litters of BB sows for this trait). No pleiotropic effect of the ESR polymorphism on average daily gain or lean meat percentage was found.
Identification of an ovulation rate QTL in cattle on BTA14 using selective DNA pooling and interval mapping1
Authors: Gonda M. G. , Arias J. A. , Shook G. E. , Kirkpatrick B. W.
Published: July 2004

Increased twinning incidence in beef cattle has the potential to improve production efficiency. However, phenotypic selection for twinning rate is difficult because of the trait's low heritability and the long time interval necessary to collect phenotypic records. Therefore, this trait and the correlated trait of ovulation rate are ideal candidates for marker‐assisted selection. The objective of this study was to complete a genome‐wide search for ovulation rate quantitative trait loci (QTL) in two related sire families. The families (paternal halfsib sires 839802 and 839803) were from a population of cattle selected for ovulation rate at the USDA Meat Animal Research Center, Clay Center, Nebraska. Putative ovulation rate QTL have previously been identified in the 839802 family on chromosomes 7 and 19; however, marker coverage in the original scan was not complete. This study fills the gaps in marker coverage of the earlier study by adding approximately 60 informative microsatellites to each sire family. Each family was genotyped using selective DNA pooling. Sons and daughters were included in either the high or low pool based on their estimated breeding value deviations from the mid‐parent average (EBVMD) for ovulation rate. Approximately 40% (839802) and 26% (839803) of available progeny comprised the high and low pools combined. Pooled typing revealed possible associations (nominal P < 0.05) between ovulation rate and marker genotype for 11 and 15 microsatellites in the 839802 and 839803 families, respectively. Subsequent interval mapping strengthened support for the presence of an ovulation rate QTL on BTA14 (chromosome‐wise P < 0.02).