bluuebunny/crossref_metadata_2025 huggingface dataset:
Cell cycle time and rate of entry of cells into mitosis in the small intestine of young rats
Authors: Wille A. C. M. , Oliveira F. A. , Soares M. A. M. , Gomes J. R.
Published: March 2004

Abstract.  Cell cycle time (TC) and the rate of entry of cells into mitosis (rM) in the jejunum and duodenum of young rats were investigated using the stathmokinetic method. The cell cycle times in the jejunum were 24.3 and 28.3 h in light and dark periods, respectively. Cell cycle times in the duodenum were 17.1 and 21.5 h in light and dark periods, respectively. Rates of entry of cells into mitosis in the jejunum were 1.2 and 1.1 cells/cell/h in light and dark periods and rates of entry of cells into mitosis in the duodenum were 1.4 and 1.8 cells/cell/h in light and dark periods, respectively. Although these changes to cell cycle time values are not statistically significant, the variation between the two periods should be considered in relation to its possible biological effects.
Oral administration of bovine whey proteins to mice elicits opposing immunoregulatory responses and is adjuvant dependent
Authors: AFUWAPE A O , TURNER M W , STROBEL S
Published: March 2004

Most studies investigating the induction of oral tolerance (OT) use purified proteins such as ovalbumin (OVA), bovine serum albumin (BSA) and beta-lactoglobulin (β-LG). Little information is available regarding the induction of OT to a protein mixture, e.g. cow's milk. In this study we compared the regulatory mechanisms induced after the oral administration of a whey protein concentrate (WP) derived from cow's milk following immunization with two different adjuvants, complete Freund's adjuvant (CFA) and alum. OVA was used as a control antigen. Animals were given a single feed of these proteins at an equivalent dose of 1 mg/g body weight before they were immunized seven days later with the antigen in Freund's adjuvant or alum. Delayed type hypersensitivity (DTH) responses were suppressed by both a feed of WP and OVA after immunization with CFA. However, only OVA feeding suppressed antigen specific IgG responses. In an attempt to investigate whether WP would tolerize the more susceptible IgE responses, alum immunization replaced CFA as the adjuvant used for systemic immunizations. WP, after a single feed, significantly primed for DTH and IgE responses indicating oral sensitization to WP. In contrast, OVA suppressed DTH, IgE and IgG responses. Antigen specific proliferation of mononuclear cells was suppressed in mice fed OVA, but primed in those fed with WP. In addition cells taken from sensitized mice fed WP up-regulated levels of specific interleukin (IL) -4, -10 and -12 in vitro whereas these cytokines were suppressed in cultures from tolerant WP fed mice. Global suppression was obtained in cultures from tolerant OVA fed mice. TGF-β was not detected in draining PLN cell cultures of either tolerant or sensitized mice. These data suggest that a whey protein mixture induces divergent responses following immunization with either CFA or alum despite being fed at an identical dose. We suggest that that the choice of the adjuvant may determine the immunoregulatory outcome and this is also reflected by the systemic cytokine profile.
Inhibition of stem cell factor reduces pulmonary cytokine levels during allergic airway responses
Authors: BERLIN A A , LINCOLN P , TOMKINSON A , LUKACS N W
Published: March 2004

Stem cell factor (SCF) has a significant role in the inflammation and activation of allergic airway responses. When monoclonal anti-SCF was administered intratracheally during allergen challenge there was a significant alteration of eosinophil accumulation and airway hyperreactivity (AHR). Anti-SCF treatment also attenuated pulmonary cytokine and chemokine levels. In particular, there was an antibody dose-dependent decrease in interleukin (IL)-5 and tumour necrosis factor (TNF)-α. There was also a significant reduction of CCL2 and CCL5, which correlated with the reduction in AHR. Mice treated with anti-SCF demonstrated a significant decrease in pulmonary gob-5 gene expression, which has been shown to correlate to goblet cell hyperplasia/metaplasia relating to airway mucus production. Blocking SCF-mediated activation within the airway using a monoclonal antibody indicates that this cytokine may represent a viable target for therapeutic intervention that could affect multiple aspects of allergen-induced immunopathology.
Regulation of complement C3 and C4 synthesis in human peritoneal mesothelial cells by peritoneal dialysis fluid
Authors: TANG S , LEUNG J C K , CHAN L Y Y , TSANG A W L , CHEN C X R , ZHOU W , LAI K N , SACKS S H
Published: March 2004

Although complement is activated in the peritoneal cavity during chronic peritoneal dialysis (PD), little is known about its role in peritoneal defence and injury related to long-term PD. We examined the impact of glucose and commercial peritoneal dialysis solutions on complement expression in HPMCs obtained by primary culture from omental tissues of consented patients undergoing elective abdominal surgery. Constitutive expression of C3 and C4 mRNA in HPMCs was up-regulated upon exposure to 75 mm glucose in a time-dependent manner. C3 and C4 protein was secreted in both apical and basolateral directions. Glucose doses beyond 100 mm markedly down-regulated C3 and C4 expression, and stimulated LDH release dose-dependently. Such cytotoxic effects were attenuated using equivalent doses of mannitol instead of glucose. Treatment with conventional lactate-buffered dialysis solution gave rise to down-regulation of C3 and C4 expression, and heightened LDH release in HPMCs. These effects correlated with the glucose strength of the solution, persisted despite replacement with a bicarbonate-buffered solution, aggravated by glycated albumin, and were partially abrogated by supplementation with 10% fetal bovine serum in the culture system. Our findings suggest that the artificial conditions imposed by PD lead to alterations in local complement synthesis that have implications for the role of the peritoneal mesothelium in both inflammation and defence.
Early deficit of lymphocytes in Wiskott–Aldrich syndrome: possible role of WASP in human lymphocyte maturation
Authors: PARK J Y , KOB M , PRODEUS A P , ROSEN F S , SHCHERBINA A , REMOLD-O'DONNELL E
Published: March 2004

Wiskott–Aldrich syndrome (WAS) is an X-linked platelet/immunodeficiency disease. The affected gene encodes WASP, a multidomain protein that regulates cytoskeletal assembly in blood cells. Patients have recurring infections, and their lymphocytes exhibit deficient proliferative responses in vitro. We report an evaluation of peripheral blood lymphocytes of 27 WAS patients, aged one month to 55 years. Whereas NK cells were normal, a significant deficit of T and B lymphocytes was observed. The number of lymphocytes was already decreased in infant patients, suggesting deficient output. Both CD4 and CD8 T lymphocytes were affected; the decrease was most pronounced for naïve T cells. Naïve CD4 lymphocytes of patients showed normal expression of Bcl-2, and Ki-67, and normal survival in vitro, suggesting that their in vivo survival and proliferation are normal. The collective data suggest that the patients’ lymphocyte deficit results from deficient output, likely due to abnormal lymphocyte maturation in the thymus and bone marrow. We propose that WASP plays an important role not only in the function of mature T lymphocytes, but also in the maturation of human T and B lymphocytes and that impaired lymphocyte maturation is central to the aetiology of WAS immunodeficiency.
Reduced ratio of protectiveversusproinflammatory cytokine responses to commensal bacteria in HLA-B27 transgenic rats
Authors: DIELEMAN L A , HOENTJEN F , QIAN B-F , SPRENGERS D , TJWA E , TORRES M F , TORRICE C D , SARTOR R B , TONKONOGY S L
Published: March 2004

Germ-free HLA-B27 transgenic (TG) rats do not develop colitis, but colonization with specific pathogen-free (SPF) bacteria induces colitis accompanied by immune activation. To study host-dependent immune responses to commensal caecal bacteria we investigated cytokine profiles in mesenteric lymph node (MLN) cells from HLA-B27 TG versus nontransgenic (non-TG) littermates after in vitro stimulation with caecal bacterial lysates (CBL). Supernatants from CBL-stimulated unseparated T- or B- cell-depleted MLN cells from HLA-B27 TG and non-TG littermates were analysed for IFN-γ, IL-12, TNF, IL-10 and TGF-β production. Our results show that unfractionated TG MLN cells stimulated with CBL produced more IFN-γ, IL-12 and TNF than did non-TG MLN cells. In contrast, CBL-stimulated non-TG MLN cells produced more IL-10 and TGF-β. T cell depletion abolished IFN-γ and decreased IL-12 production, but did not affect IL-10 and TGF-β production. Conversely, neither IL-10 nor TGF-β was produced in cultures of B cell-depleted MLN. In addition, CD4+ T cells enriched from MLN of HLA-B27 TG but not from non-TG rats produced IFN-γ when cocultured with CBL-pulsed antigen presenting cells from non-TG rats. Interestingly, IL-10 and TGF-β, but not IFN-γ, IL-12 and TNF were produced by MLN cells from germ-free TG rats. These results indicate that the colitis that develops in SPF HLA-B27 TG rats is accompanied by activation of IFN-γ-producing CD4+ T cells that respond to commensal bacteria. However, B cell cytokine production in response to components of commensal intestinal microorganisms occurs in the absence of intestinal inflammation.
The membrane attack complex, C5b-9, up regulates collagen gene expression in renal tubular epithelial cells
Authors: ABE K , LI K , SACKS S H , SHEERIN N S
Published: March 2004

Evidence suggesting a direct role for proteinuria in the pathogenesis of renal tubulointerstitial fibrosis is accumulating. However the mechanism by which proteinuria leads to injury is unknown. In proteinuric states complement proteins are filtered through the glomerulus and could contribute to the tubular damage. The aim of this study was to investigate the role of complement activation in the progression of interstitial fibrosis. To determine whether complement activation may be responsible for the pro-fibrotic response that occurs in the tubulointerstitial compartment we stimulated primary cultures of proximal tubular epithelial cells with membrane attack complex, C5b-9. This led to increased mRNA concentrations of both collagen type IV and its intracellular chaperone, Heat Shock Protein 47 (HSP47). To determine whether this occurred in vivo Adriamycin was used to induce proteinuria in female Balb/c mice. The expression of collagen type IV and HSP47 was increased in proteinuric mice compared to control mice. In proteinuric mouse kidney, C3 was deposited at sites of tubulointerstitial injury and there was a relationship between C3 deposition and immunochemical staining for collagen type IV and HSP47. In situ hybridization suggested that the renal tubular epithelium was actively expressing HSP47 mRNA and, by implication, excess collagen. These observations support the hypothesis that complement activation on tubular epithelial cells can directly increase the pro-fibrotic process associated with tubulointerstitial damage.
Differential effect of calcium phosphate and calcium pyrophosphate on binding of matrix metalloproteinases to fibrin: comparison to a fibrin-binding protease from inflammatory joint fluids
Authors: MAKOWSKI G S , RAMSBY M L
Published: March 2004

The ability of calcium phosphate (CaP) and calcium pyrophosphate (CaPPi) to mediate matrix metalloproteinase-2 and -9 (MMP-2 and MMP-9) binding to fibrin was evaluated. Substrate gel electrophoresis (gelatin zymography) revealed that CaP bound MMP-2 and MMP-9, forming a high molecular weight aggregate with lowered electrophoretic mobility. Formation of the CaP : MMP aggregate was necessary for fibrin binding. In contrast, CaPPi did not aggregate MMPs and did not promote uptake of MMPs into fibrin. Scatchard analysis (Ca/P ratio) revealed that CaPPi (1·96) was chemically similar to calcium pyrophosphate dihydrate (2·00) compared to amorphous CaP (1·50) or crystalline CaP, hydroxyapatite (1·66). MMP : CaP interaction appeared to be electrostatic in nature as high salt concentration (NaCl > 150 mm) reduced binding. In contrast, two non-ionic detergents (Brij-35 and Tween-20) did not prevent MMP : CaP binding. MMP : CaP interaction did not involve the C-terminal MMP region because the specific tissue inhibitor of metalloproteinases (TIMPs) also did not block MMP : CaP interaction and fibrin binding. Although MMP : CaP binding could be decreased with albumin, this effect appeared non-specific due to the high albumin concentration required. High albumin concentration could also partially dissociate preformed MMP : CaP complexes. Interestingly, type I and type IV collagen substantially increased MMP : fibrin-binding activity, whereas denatured collagen, gelatin, did not. Inflammatory joint fluid from five patients also demonstrated similar MMP fibrin-binding activity consistent with CaP mediation. The relevance of these findings to CaP and CaPPi in the pathogenesis of crystal arthropathies such as basic calcium phosphate (BCP) and calcium pyrophosphate dihydrate crystal disease (CPPD) is discussed.
Antibodies to TSH-receptor in thyroid autoimmune disease interact with monoclonal antibodies whose epitopes are broadly distributed on the receptor
Authors: MINICH W B , LENZNER C , MORGENTHALER N G
Published: March 2004

The hyperthyroidism of Graves’ disease (GD) is caused by TSH-receptor (TSH-R) stimulating autoantibodies (TSAb), leading to overproduction of thyroid hormones. We present evidence for TSAb interaction with three distinct regions of the TSH-R, one in immediate vicinity of the carboxy terminal serpentine. Three murine monoclonal antibodies (MoAbs 28·1, A9 and 31·7) directed to amino acids 36–40, 147–228 and 382–415 were labelled and tested for their binding to human recombinant TSH-R on solid phase. All MoAbs bound to TSH-R with a Kd of 8–12 nm and showed no competition among themselves. We tested 114 sera from euthyroid controls, 118 TBII positive sera from patients with GD (containing TSAb confirmed by bioassays), 16 TBII positive sera from patients with autoimmune thyroid disease (AIT), who were hypothyroid and had TSH blocking antibodies (TBAb), and 20 patients with AIT, who were hypothyroid but negative for all TRAb. Mid-regional MoAb A9 tracer achieved the highest sensitivity in the GD group (72·0%), whereas C-terminal MoAb 31·7 found most sera positive in the AIT group (87·5%). Surprisingly, the N-terminal MoAb 28·1 had the lowest sensitivity in the GD (10·4%) and AIT group (43·8%). Using a mixture of all three tracer MoAbs did not increase the sensitivity in the GD or AIT group, compared to the best single MoAb alone. Median inhibition of MoAb A9 was significantly (P < 0·001) higher than inhibition of MoAbs 28·1 or 31·7 in the group of GD patients but not in other groups. Almost all patient sera with positive reactivity in the MoAb tracer assays had TBII values in the higher range. However, there were many highly TBII positive sera, which did not show a displacement of the MoAb tracers. We conclude that, contrary to some reports, the binding of TSAb and TBAb to the TSH-R is not restricted to distinct and distant epitopes. The middle part of the TSH-R seems to be more relevant for TSAb binding than the N-terminal part, while a proportion of TSAb autoantibodies also binds to a C-terminal epitope of the TSH-R. The method described here is a TSH independent competitive assay for the detection of TSH-R autoantibodies.
Induction of experimental autoimmune encephalomyelitis in Dark Agouti rats without adjuvant
Authors: STOSIC-GRUJICIC S , RAMIC Z , BUMBASIREVIC V , HARHAJI L , MOSTARICA-STOJKOVIC M
Published: March 2004

Experimental autoimmune encephalomyelitis (EAE) is a well-recognized model for multiple sclerosis (MS) in humans. However, adjuvants used with encephalitogens to induce EAE produce non-specific effects interfering with the mechanisms involved in the autoimmune response to the central nervous system (CNS) tissue. It is therefore important to establish a more suitable model of EAE for analysis of autoimmune phenomena resembling those operative in MS. Here we report that EAE can be induced regularly in Dark Agouti (DA) strain of rats with spinal cord tissue without any adjuvant, as judged by both clinical and histological parameters. The incidence and severity of EAE depended on the origin of the encephalitogen, the rat versus guinea pig spinal cord homogenate being more efficient. Furthermore, EAE could be reinduced in animals which had recovered from disease that had been induced actively with encephalitogen alone, suggesting the role of adjuvant-generated non-specific mechanisms in resistance to reinduction of EAE. Thus, EAE induced in DA rats with encephalitogen alone provides a reproducible model for defining pathogenically relevant events in CNS autoimmunity devoid of the potentially misleading effects of adjuvants.
P-selectin glycoprotein ligand 1 therapy ameliorates established collagen-induced arthritis in DBA/1 mice partly through the suppression of tumour necrosis factor
Authors: SUMARIWALLA P F , MALFAIT A M , FELDMANN M
Published: March 2004

We investigated the therapeutic potential of P-selectin glycoprotein ligand (PSGL)-1 in established collagen-induced arthritis (CIA) in DBA/1 mice. PSGL-1 is the high-affinity specific ligand for P-selectin and is thus important in cell recruitment to inflammatory sites. I-316 PSGL-1 or rPSGL-1Ig fusion protein were administered to mice after the onset of clinical arthritis for 10 days, and the effect of treatment on both clinical and histopathological progression of disease was studied. It was found that both PSGL-1 biologicals effectively suppressed progression of clinical arthritis, and this was accompanied by protection against damage of joint tissues. We sought to investigate a mechanism underlying the effect of rPSGL-1Ig on the reduction of clinical arthritis. Blockade of PSGL-1/P–selectin interaction blocks recruitment of leucocytes, thus we observed a notable reduction in viable cell numbers of synoviocytes from rPSGL-1Ig treated mice. In view of this finding we suspected an effect of treatment on the production of pro-inflammatory mediators such as bioactive tumour necrosis factor-α (TNF) in synovial membrane ex vivo cell cultures. Production of TNF was reduced in arthritic mice that had been treated with rPSGL-1Ig. To further investigate the mechanism of rPSGL-1Ig, we explored the possibility that PSGL-1 might also have a direct signalling effect on TNF release from inflammatory cells. Thus synoviocyte cultures from arthritic mice were incubated with rPSGL-1Ig. A significant reduction in the spontaneous bioactive TNF release from these cultures was noted. We therefore confirmed these surprising findings using cultures of a mouse macrophage like cell line RAW 264·7, stimulated by LPS. Our results indicate that both forms of PSGL-1 have significant therapeutic effects in CIA murine model of RA. The mechanism of action involves reduced cellularity of synovium as anticipated, along with a reduction in TNF production from inflammatory cells in the synovium. The latter mechanism needs further mechanistic analysis.
Evaluation of conformational epitopes on thyroid peroxidase by antipeptide antibody binding and mutagenesis
Authors: GORA M , GARDAS A , WIKTOROWICZ W , HOBBY P , WATSON P F , WEETMAN A P , SUTTON B J , BANGA J P
Published: March 2004

Autoantibodies to thyroid peroxidase (TPO) recognize predominantly conformational epitopes, which are restricted to two distinct determinants, termed immunodominant domain region (IDR) A and B. These dominant determinants reside in the region with structural homology to myeloperoxidase (MPO)-like domain and may extend into the adjacent complement control protein (CCP) domain. We have explored the location of these determinants on the MPO-like domain of the structural model of TPO, by identifying exposed hydrophilic loops that are potential candidates for the autoantigenic sites, generating rabbit antipeptide antisera, and competing with well characterized murine monoclonal antibodies (mabs) specific for these two IDRs. We recently defined the location of IDR-B, and here report our findings on the location of IDR-A and its relationship to IDR-B, defined with a new panel of 15 antipeptide antisera. Moreover, in combination with single amino acid replacements by in vitro mutagenesis, we have defined the limits of the IDR-B region on the TPO model. The combination of antisera to peptides P12 (aa 549–563), P14 (aa 599–617) and P18 (aa 210–225) inhibited the binding of the mab specific for IDR-A (mab 2) by 75%. The same combination inhibited the binding of autoantibodies to native TPO from 67 to 94% (mean 81·5%) at autoantibody levels of 5 IU. Fabs prepared from the antipeptide IgG and pooled in this combination were also effective in competition assays, thus defining the epitopes more precisely. IDR-A was found to lie immediately adjacent to IDR-B and thus the two immunodominant epitopes form an extended patch on the surface of TPO. Finally, by single amino acid mutagenesis, we show that IDR-B extends to residue N642, thus further localizing the boundary of this autoantigenic region on the structural model.
Recombination activating genes (RAG) induce secondary Ig gene rearrangement in and subsequent apoptosis of human peripheral blood circulating B lymphocytes
Authors: NAGAFUCHI H , YOSHIKAWA H , TAKEBA Y , NARA K , MIURA K , KUROKAWA M S , SUZUKI N
Published: March 2004

Recombination activating gene (RAG) re-expression and secondary Ig gene rearrangement in mature B lymphocytes have been reported. Here, we have studied RAG expression of peripheral blood B lymphocytes in humans. Normal B cells did not express RAG1 and RAG2 spontaneously. More than a half of circulating B cells expressed RAG proteins, when activated with Staphylococcus aureus Cowan I (SAC) + IL-2. DNA binding activity of the RAG complex has been verified by a gel shift assay employing the recombination signal sequence (RSS). Secondary Ig light chain rearrangement in the RAG-expressing B cells was confirmed by linker-mediated (LM)-PCR. Highly purified surface κ+ B cells activated by SAC + IL-2 became RAG+, and thereafter they started to express λ chain mRNA. 2 colour immunofluorescence analysis disclosed that a part of the RAG+ cells derived from the purified κ+ B cells activated by SAC + IL-2 turned to λ+ phenotype in vitro. Similarly, apoptosis induction was observed in a part of the RAG+ B cells. Our study suggests that a majority of peripheral blood B cells re-expresses RAG and the RAG+ B lymphocytes could be eliminated from the B cell repertoire either by changing Ag receptor specificity due to secondary rearrangement or by apoptosis induction. Thus, RAG expression of mature B cells in peripheral blood would contribute to not only receptor revision for further diversification of B cell repertoire but in some cases (or in some B cell subsets) to prevention or induction of autoAb responses at this differentiation stage in humans.
Unravelling the genetic complexity of autoimmune thyroid disease: HLA, CTLA-4 and beyond
Authors: SIMMONDS M J , GOUGH S C L
Published: March 2004

The autoimmune thyroid diseases (AITDs) including Graves’ disease (GD) and autoimmune hypothyroidism (AIH) are the commonest of the autoimmune conditions affecting 2–5% of the western population. Twin studies have clearly demonstrated that AITDs are caused by a combination of both environmental and genetic factors. Association of the HLA class II region with AITD has been documented for over 20 years now, but the primary aetiological variant in this region remains unknown. More recently the CTLA-4 gene region has been identified as the second locus conferring susceptibility to AITD. In contrast to HLA, a polymorphism of the CTLA-4 gene, which encodes an important negative regulator of the immune system, has been identified as a candidate for a primary determinant for AITD. A large number of candidate gene and genome wide linkage studies have been involved in the search for the elusive ‘third’ locus. The thyroglobulin (Tg) gene in humans maps to chromosome 8q, which has been linked in family studies to AITD. A number of association studies in humans and the mouse model for AITD are beginning to implicate the Tg gene although convincing evidence for a primary causative role is still needed. The establishment of large DNA disease resources along with more detailed genetic maps and the development of faster, more effective, high throughput genotyping and sequencing methods, provides some sense of optimism that novel loci will be identified in the near future and the complex aetiology of AITD will be further unraveled.
Pedagogical underpinnings of computer‐based learning
Authors: Adams Audrey M.
Published: March 2004

Background.  E‐learning is becoming increasingly incorporated into educational programmes. Digital materials usually require a lot of investment in terms of time, money and human resources. With advances in technology, delivery of content has much improved in terms of multimedia elements. However, often only low‐level learning is achieved as a result of using these materials.

Aim.  The purpose of this article is to give a comprehensive overview of some of the most important issues to consider when incorporating e‐learning into educational programmes.

Discussion.  Computer‐based learning has three components: hardware, software and ‘underware’, the pedagogy that underpins its development. The latter is the most important, as the approach adopted will influence the creation of computer‐based learning materials and determine the way in which students engage with subject matter. Teachers are responsible for the quality of their courses and have a vital role in helping to develop the most appropriate electronic learning activities that will facilitate students to acquire the knowledge and skills necessary for clinical practice. Therefore, they need to have an awareness of what contributes to educationally effective, computer‐based learning materials.
Older people with depression: pilot study
Authors: Minardi Henry A. , Blanchard Martin
Published: March 2004

Background.  Although a substantial number of studies have identified the prevalence of depression experienced by older adults living in the community and the factors that may precipitate this depression, few have focussed on depression in older people who attend day centres. Consequently, there is no literature suggesting how depression in this population may be psychologically managed.

Aims.  The aim of this paper is to present the results of a pilot study investigating the incidence of depression in people attending a London Age Concern day centre, and its associations with perceptions of handicap, loneliness, social support networks, satisfaction with social support, and satisfaction with life.

Methods.  A quasi‐experimental, cross‐sectional pilot study was conducted with attendees at a London Age Concern day centre between November 1998 and January 1999. The day centre manager selected a convenience sample of 24 of 156 participants. Inclusion criteria were that participants must attend the day centre regularly and be over or equal to 65 years of age. The exclusion criterion was known psychiatric illness. Participants were interviewed using the Geriatric Mental State‐Automated Geriatric Examination for Computer Assisted Taxonomy, which identifies caseness of psychopathology and includes a loneliness subscale, and completed the Satisfaction with Life Scale, Social Support Questionnaire and London Handicap Scale.

Results.  There was a higher level of depression in this day centre setting than that found in other community‐based studies. There were the expected significant associations between depression and loneliness, and depression and satisfaction with life. However, unexpectedly, there were no associations between depression and social support, or depression and handicap.

Conclusions.  The study has several limitations. However, the high level of depression and significant relationships between depression, loneliness and satisfaction with life may indicate that the social function of day centres is not meeting expectations. This suggests that there could be a role for community mental health nurses in offering psychosocial interventions to alleviate some of the distress experienced.
How delivery ward staff exercise power over women in communication
Authors: Sinivaara Maria , Suominen Tarja , Routasalo Pirkko , Hupli Maija
Published: March 2004

Background.  Power is a central element in all social interactions. In order to act appropriately in different care situations, it is important to understand the meaning of power.

Aim.  This paper reports a study whose aim was to describe delivery ward staff opinions about the exercise of power over women in communications.

Methods.  Data were collected in 2000 using a questionnaire developed for the study with a convenience sample of 155 midwifery and nursing professionals in delivery wards in four Finnish hospitals.

Results.  The results showed that in respondents’ opinions they worked for women's good and aimed at relationships based on a sense of equality and individuality. Negative power was used in situations where women's well‐being was threatened. The power used was verbal or non‐verbal.

Conclusions.  The exercise of power is not only negative in nature. There are times, such as in life‐threatening situations, when it is crucial that the professional takes control and makes decisions immediately, and in that sense exercises power.
A systematic review to ascertain whether the standard needle is more effective than a longer or wider needle in reducing the incidence of local reaction in children receiving primary immunization
Authors: Davenport Julie M.
Published: March 2004

Context.  Millions of doses of vaccines are given to children within the United Kingdom (UK) each year, yet there is conflicting national guidance on the length of needle to use. Prefilled syringes come supplied with 25 Gauge 16 millimetres long needles.

Objectives.  A systematic review was completed to answer the clinical question of whether the standard needle is more effective than a longer and/or wider needle in reducing local reactions in children receiving primary immunizations.

Methods.  Data sources: A comprehensive search strategy, including searching of electronic databases, hand searching, reference/citation searching, contacting researchers in the field and searching for unpublished literature. Study selection: Selection of studies was guided by the clinical question, the inclusion and exclusion criteria being applied by two people independently. Study quality assessment: An established quality checklist was used. Critical analysis of the quality of the components of the studies is included in the discussion. Data extraction: Two people independently applied a data extraction form, designed for the study.

Results.  (1) Rev Man (a Windows‐based software package used by the Cochrane Collaboration) was used to perform a meta‐analysis on the binary results of two small studies using Relative Risk as the summary statistic 16 mm vs. 25 mm needle. (2) The pooled Relative Risk for redness was 2·52 [95% confidence interval (CI) fixed 1·70–3·72] P < 0·00001. (3) The pooled Relative Risk for swelling was 2·31 (95% CI fixed 1·55–3·43) P < 0·00003.

Conclusion.  (1) The results are statistically very highly significant and favour the use of the 25 millimetre long needle. (2) No studies were identified to separate the effect of the gauge of the needle used, but the discussion shows that needle length is the most important variable. (3) The results should be incorporated into any future guidelines on vaccine administration in the UK.
Practical application of the Delphi technique in a bicultural mental health nursing study in New Zealand
Authors: Hardy Derrylea J. , O'Brien Anthony P. , Gaskin Cadeyrn J. , O'Brien Anthony J. , Morrison‐Ngatai Erina , Skews Georgina , Ryan Tom , McNulty Neil
Published: March 2004

Background.  Numerous studies have employed the Delphi technique to seek expert opinion about aspects of clinical practice. When researching literature on the Delphi technique, however, we discovered discrepancies in its application, and a lack of detail when reporting design, administration, and analysis methods. Such lack of specificity hinders the replicability and assessment of the clinical and cultural validity and reliability of Delphi studies.

Aim.  The aim of this paper is to detail the practical application of the Delphi technique as a culturally and clinically valid means of accessing expert opinion on the importance of clinical criteria.

Methods.  Reference is made to a bicultural New Zealand mental health nursing clinical indicator study that employed a three‐round reactive Delphi survey. Equal proportions of Maori and non‐Maori nurses (n = 20) and consumers (n = 10) rated the importance of 91 clinical indicator statements for the achievement of professional practice standards. Additional statements (n = 21) suggested by Delphi participants in round 1 were included in subsequent rounds. In round 2, participants explained the rating they applied to statements that had not reached consensus in round 1, and summarized responses were provided to participants in round 3. Consensus was considered to have been achieved if 85% of round 3 ratings lay within a 2‐point bracket on the 5‐point Likert‐scale overall, or in one of the Maori nurse, non‐Maori nurse, or consumer groups. A mean rating of 4·5 after round 3 was set as the importance threshold.

Findings.  Consensus occurred overall on 75 statements, and within groups on another 24. Most statements (n = 86) reached the importance benchmark.

Conclusions.  When rigorous methods of participant selection, group composition, participant feedback, and determination of consensus and importance are employed, the Delphi technique is a reliable, cost‐effective means of obtaining and prioritizing experts judgements.
Treatment protocols for demodicosis: an evidence‐based review
Authors: Mueller Ralf S.
Published: March 2004

Abstract Publications discussing the treatment of demodicosis in the dog and cat are reviewed. Based on the evidence in the literature, amitraz rinses at 0.025–0.06% every 7–14 days, and oral daily ivermectin at 300 µg kg−1, milbemycin at 2 mg kg−1and moxidectin at 400 µg kg−1, respectively, can all be recommended for the treatment of generalized canine demodicosis. Ivermectin and moxidectin should be initiated at lower doses and patients monitored for possible adverse effects during therapy. In cats, 2% lime sulfur dips and amitraz rinses at 0.0125–0.025% have been used successfully.
Treatment of dermatophytosis in dogs and cats: review of published studies
Authors: Moriello Karen A.
Published: March 2004

Abstract The recent literature on the treatment of dermatophytosis in dogs and cats was reviewed. Based uponin vitrostudies using isolated infected hairs and controlled or fieldin vivostudies, the following topical treatments were consistently found to be antifungal (i.e. antidermatophyte): lime sulfur (1:16), 0.2% enilconazole rinses, and a combined 2% miconazole/chlorhexidine shampoo. Animals or hairs were either bathed or rinsed once or twice weekly. Itraconazole, griseofulvin and terbinafine were evaluated in controlled or field studies, most commonly involving cats. Griseofulvin (50 mg kg−1) was reported to cure infected animals in 41–70 days. Itraconazole (10 mg kg−1once daily or in a combined daily/pulse therapy 10 mg kg−1once daily for 28 days and then week on/week off) was reported to cure infected animals in 56–70 days. Low‐dose itraconazole (1.5–3.0 mg kg−1) in 15‐day cycles required 1–3 cycles (15–45 days). Various doses of terbinafine (5–40 mg kg−1) were reportedly used to treat dogs or cats. The higher doses of terbinafine (> 20 mg kg−1) were required to achieve a mycological cure; the number of treatment days to cure varied from 21 to > 126 days. Lufenuron was reported anecdotally to be an effective cure, however, this was not substantiated in controlled studies. Finally, fungal vaccines were not found to be effective against challenge exposure, however, there is evidence that they may be useful in treatment protocols.
Current trends in the treatment of Sarcoptes, Cheyletiella and Otodectes mite infestations in dogs and cats
Authors: Curtis Cathy F.
Published: March 2004

Abstract  For a number of reasons, several of the more ‘traditional’ ectoparasiticides in the small animal veterinarian's armoury have been withdrawn over the past few years. New, safer products which are long‐acting and easier to apply than the conventional dips, rinses and aerosol sprays of the past have replaced them. However, relatively few such novel acaricidal preparations have become commercially available. Consequently, practitioners and researchers frequently experiment with the drugs they have at their disposal to assess their efficacy against a variety of target acarids when used at different dosages and/or via different routes of administration, compared with those recommended by the manufacturer. This paper reviews the anecdotal and peer‐reviewed reports describing the use of modern acaricides in dogs and cats that have recently appeared in the veterinary literature. It should be stressed, however, that no medicine should be prescribed for extra‐label use without the informed consent of the owner.
Golgi Inheritance Under a Block of Anterograde and Retrograde Traffic
Authors: Nizak Clément , Sougrat Rachid , Jollivet Florence , Rambourg Alain , Goud Bruno , Perez Franck
Published: March 2004

In mitosis, the Golgi complex is inherited following its dispersion, equal partitioning and reformation in each daughter cell. The state of Golgi membranes during mitosis is controversial, and the role of Golgi‐intersecting traffic in Golgi inheritance is unclear. We have used brefeldin A (BFA) to perturb Golgi‐intersecting membrane traffic at different stages of the cell cycle and followed by live cell imaging the fate of Golgi membranes in those conditions. We observed that addition of the drug on cells in prometaphase prevents mitotic Golgi dispersion. Under continuous treatment, Golgi fragments persist throughout mitosis and accumulate in a Golgi‐like structure at the end of mitosis. This structure localizes at microtubule minus ends and contains all classes of Golgi markers, but is not accessible to cargo from the endoplasmic reticulum or the plasma membrane because of the continuous BFA traffic block. However, it contains preaccumulated cargo, and intermixes with the reforming Golgi upon BFA washout. This structure also forms when BFA is added during metaphase, when the Golgi is not discernible by light microscopy. Together the data indicate that independent Golgi fragments that contain all classes of Golgi markers (and that can be isolated from other organelles by blocking anterograde and retrograde Golgi‐intersecting traffic) persist throughout mitosis.
Point mutations associated with insecticide resistance in the Drosophila cytochrome P450 Cyp6a2 enable DDT metabolism
Authors: Amichot Marcel , Tarès Sophie , Brun‐Barale Alexandra , Arthaud Laury , Bride Jean‐Marc , Bergé Jean‐Baptiste
Published: March 2004

Three point mutations R335S, L336V and V476L, distinguish the sequence of a cytochrome P450 CYP6A2 variant assumed to be responsible for 1,1,1‐trichloro‐2,2‐bis‐(4′‐chlorophenyl)ethane (DDT) resistance in the RDDTR strain of Drosophila melanogaster. To determine the impact of each mutation on the function of CYP6A2, the wild‐type enzyme (CYP6A2wt) of Cyp6a2 was expressed in Escherichia coli as well as three variants carrying a single mutation, the double mutant CYP6A2vSV and the triple mutant CYP6A2vSVL. All CYP6A2 variants were less stable than the CYP6A2wt protein. Two activities enhanced in the RDDTR strain were measured with all recombinant proteins, namely testosterone hydroxylation and DDT metabolism. Testosterone was hydroxylated at the 2β position with little quantitative variation among the variants. In contrast, metabolism of DDT was strongly affected by the mutations. The CYP6A2vSVL enzyme had an enhanced metabolism of DDT, producing dicofol, dichlorodiphenyldichloroethane and dichlorodiphenyl acetic acid. The apparent affinity of the enzymes CYP6A2wt and CYP6A2vSVL for DDT and testosterone was not significantly different as revealed by the type I difference spectra. Sequence alignments with CYP102A1 provided clues to the positions of the amino acids mutated in CYP6A2. These mutations were found spatially clustered in the vicinity of the distal end of helix I relative to the substrate recognition valley. Thus this area, including helix J, is important for the structure and activity of CYP6A2. Furthermore, we show here that point mutations in a cytochrome P450 can have a prominent role in insecticide resistance.
The C‐terminal domain of Escherichia coli Hfq increases the stability of the hexamer
Authors: Arluison Véronique , Folichon Marc , Marco Sergio , Derreumaux Philippe , Pellegrini Olivier , Seguin Jérôme , Hajnsdorf Eliane , Regnier Philippe
Published: March 2004

The Hfq (Host factor 1) polypeptide is a nucleic acid binding protein involved in the synthesis of many polypeptides. Hfq particularly affects the translation and the stability of several RNAs. In an earlier study, the use of fold recognition methods allowed us to detect a relationship between Escherichia coli Hfq and the Sm topology. This topology was further validated by a series of biophysical studies and the Hfq structure was modelled on an Sm protein. Hfq forms a β‐sheet ring‐shaped hexamer. As our previous study predicted a large number of alternative conformations for the C‐terminal region, we have determined whether the last 19 C‐terminal residues are necessary for protein function. We find that the C‐terminal truncated protein is fully capable of binding a polyadenylated RNA (Kd of 120 pm vs. 50 pm for full‐length Hfq). This result shows that the functional core of E. coli Hfq resides in residues 1–70 and confirms previous genetic studies. Using equilibrium unfolding studies, however, we find that full‐length Hfq is 1.8 kcal·mol−1 more stable than its truncated variant. Electron microscopy analysis of both truncated and full‐length proteins indicates a structural rearrangement between the subunits upon truncation. This conformational change is coupled to a reduction in β‐strand content, as determined by Fourier transform infra‐red. On the basis of these results, we propose that the C‐terminal domain could protect the interface between the subunits and stabilize the hexameric Hfq structure. The origin of this C‐terminal domain is also discussed.
Heterogeneity of homologously expressed Hypocrea jecorina (Trichoderma reesei) Cel7B catalytic module
Authors: Eriksson Torny , Stals Ingeborg , Collén Anna , Tjerneld Folke , Claeyssens Marc , Stålbrand Henrik , Brumer Harry
Published: March 2004

The catalytic module of Hypocrea jecorina (previously Trichoderma reesei) Cel7B was homologously expressed by transformation of strain QM9414. Post‐translational modifications in purified Cel7B preparations were analysed by enzymatic digestions, high performance chromatography, mass spectrometry and site‐directed mutagenesis. Of the five potential sites found in the wild‐type enzyme, only Asn56 and Asn182 were found to be N‐glycosylated. GlcNAc2Man5 was identified as the predominant N‐glycan, although lesser amounts of GlcNAc2Man7 and glycans carrying a mannophosphodiester bond were also detected. Repartition of neutral and charged glycan structures over the two glycosylation sites mainly accounts for the observed microheterogeneity of the protein. However, partial deamidation of Asn259 and a partially occupied O‐glycosylation site give rise to further complexity in enzyme preparations.
SUT2 is a novel multicopy suppressor of low activity of the cAMP/protein kinase A pathway in yeast
Authors: Rützler Michael , Reissaus André , Budzowska Magdalena , Bandlow Wolfhard
Published: March 2004

SUT2 was found in a screen for multicopy suppressors of the synthetic slow growth phenotype of a Δras2Δgpa2 double deletion mutant. It failed, however, to cure the lethal phenotype of a Δras1Δras2 mutant suggesting that it acts upstream of Ras or in a parallel pathway. By testing cAMP‐dependent reactions including the accumulation of storage carbohydrates, pseudohyphal differentiation, entry of meiosis as well as the measurement of FLO11 reporter activity we show that Sut2p modulates the activity of protein kinase A (PKA). Additionally, we demonstrate that cellular levels of Ras2p are affected by Sut2p and that Sut2‐GFPp accumulates significantly in the nucleus. Based on the observed influence of high SUT2 gene dosage on PKA activity as well as Sut2p's homology to the presumptive transcription factor Sut1p, we suggest that Sut2p contributes to regulation of PKA activity at the level of transcription.
Disruption of the interaction between the Rieske iron–sulfur protein and cytochrome b in the yeast bc1 complex owing to a human disease‐associated mutation within cytochrome b
Authors: Fisher Nicholas , Bourges Ingrid , Hill Philip , Brasseur Gael , Meunier Brigitte
Published: March 2004

The mitochondrial cytochrome b missense mutation, G167E, has been reported in a patient with cardiomyopathy. The residue G167 is located in an extramembranous helix close to the hinge region of the iron–sulfur protein. In order to characterize the effects of the mutation on the structure and function of the bc1 complex, we introduced G167E into the highly similar yeast cytochrome b. The mutation had a severe effect on the respiratory function, with the activity of the bc1 complex decreased to a few per cent of the wild type. Analysis of the enzyme activity indicated that the mutation affected its stability, which could be the result of an altered binding of the iron–sulfur protein on the complex. G167E had no major effect on the interaction between the iron–sulfur protein headgroup and the quinol oxidation site, as judged by the electron paramagnetic resonance signal, and only a minor effect on the rate of cytochrome b reduction, but it severely reduced the rate of cytochrome c1 reduction. This suggested that the mutation G167E could hinder the movement of the iron–sulfur protein, probably by distorting the structure of the hinge region. The function of bc1 was partially restored by mutations (W164L and W166L) located close to the primary change, which reduced the steric hindrance caused by G167E. Taken together, these observations suggest that the protein–protein interaction between the n‐sulfur protein hinge region and the cytochrome b extramembranous cd2 helix is important for maintaining the structure of the hinge region and, by consequence, the movement of the headgroup and the integrity of the enzyme.