Streptococcus mutans glucan-binding protein-A affects Streptococcus gordonii biofilm architecture

The glucan-binding protein-A (GbpA) of Streptococcus mutans has been shown to contribute to the architecture of glucan-dependent biofilms formed by this species and influence virulence in a rat model. As S. mutans synthesizes multiple glucosyltransferases and nonglucosyltransferase glucan-binding proteins (GBPs), it is possible that there is functional redundancy that overshadows the full extent of GbpA contributions to S. mutans biology. Glucan-associated properties such as adhesion, aggregation, and biofilm formation were examined independently of other S. mutans GBPs by cloning the gbpA gene into a heterologous host, Streptococcus gordonii, and derivatives with altered or diminished glucosyltransferase activity. The presence of GbpA did not alter dextran-dependent aggregation nor the initial sucrose-dependent adhesion of S. gordonii. However, expression of GbpA altered the biofilm formed by wild-type S. gordonii as well as the biofilm formed by strain CH107 that produced primarily alpha-1,6-linked glucan. Expression of gbpA did not alter the biofilm formed by strain DS512, which produced significantly lower quantities of parental glucan. These data are consistent with a role for GbpA in facilitating the development of biofilms that harbor taller microcolonies via binding to alpha-1,6-linkages within glucan. The magnitude of the GbpA effect appears to be dependent on the quantity and linkage of available glucan.     

Clinical validation of an immunoaffinity LC-MS/MS assay for the quantification of a collagen type II neoepitope peptide: A biomarker of matrix metalloproteinase activity and osteoarthritis in human urine

The degradation of type II collagen has been associated with the pathology of osteoarthritis (OA). Matrix metalloproteinases (MMPs) are enzymes that are responsible for catalyzing the degradation of collagen and, therefore, are pursued as potential targets for the treatment of OA. Collagen-derived peptides identified as a reflection of in vivo MMP activity have been investigated as target biomarkers of MMP activity as well as potential biomarkers of OA disease state and/or progression. An immunoaffinity liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay developed for the quantification of the most abundant urinary type II collagen neoepitope (uTIINE) peptide, a 45-mer with 5 HO-proline residues resulting from MMP-13-catalyzed degradation, was validated for clinical use. Validation experiments were designed with attention to specific challenges related to quantification of endogenous analytes. The validated method is sensitive, selective, accurate (<15% relative error) and precise (<15% coefficient of variation) over a linear range of 0.156-7.50 ng/ml. Sample stability and inter- and intrasubject variability were evaluated in the urine of normal and OA populations. The method was applied to analyze human urine samples from clinical studies investigating the utility of uTIINE as a potential biomarker for OA.     

The eVALuate study: two parallel randomised trials,one comparing laparoscopic with abdominal hysterectomy,the other comparing laparoscopic with vaginal hysterectomy

Objective To compare the effects of laparoscopic hysterectomy and abdominal hysterectomy in the abdominal trial, and laparoscopic hysterectomy and vaginal hysterectomy in the vaginal trial.Design Two parallel, multicentre, randomised trials.Setting 28 UK centres and two South African centres.Participants 1380 women were recruited; 1346 had surgery; 937 were followed up at one year.Primary outcome Rate of major complications.Results In the abdominal trial laparoscopic hysterectomy was associated with a higher rate of major complications than abdominal hysterectomy (11.1% v 6.2%, P = 0.02; difference 4.9%, 95% confidence interval 0.9% to 9.1%) and the number needed to treat to harm was 20. Laparoscopic hysterectomy also took longer to perform (84 minutes v 50 minutes) but was less painful (visual analogue scale 3.51 v 3.88, P = 0.01) and resulted in a shorter stay in hospital after the operation (3 days v 4 days). Six weeks after the operation, laparoscopic hysterectomy was associated with less pain and better quality of life than abdominal hysterectomy (SF-12, body image scale, and sexual activity questionnaires).In the vaginal trial we found no evidence of a difference in major complication rates between laparoscopic hysterectomy and vaginal hysterectomy (9.8% v 9.5%, P = 0.92; difference 0.3%, -5.2% to 5.8%), and the number needed to treat to harm was 333. We found no evidence of other differences between laparoscopic hysterectomy and vaginal hysterectomy except that laparoscopic hysterectomy took longer to perform (72 minutes v 39 minutes) and was associated with a higher rate of detecting unexpected pathology (16.4% v 4.8%, P = < 0.01). However, this trial was underpowered.Conclusions Laparoscopic hysterectomy was associated with a significantly higher rate of major complications than abdominal hysterectomy. It also took longer to perform but was associated with less pain, quicker recovery, and better short term quality of life. The trial comparing vaginal hysterectomy with laparoscopic hysterectomy was underpowered and is inconclusive on the rate of major complications; however, vaginal hysterectomy took less time.     

Active hexose correlated compound enhances resistance to Klebsiella pneumoniae infection in mice in the hindlimb-unloading model of spaceflight conditions.

Previous studies have demonstrated that resistance to infection is decreased in Swiss Webster female mice maintained in the hindlimb-unloading model (Aviles H, Belay T, Fountain K, Vance M, and Sonnenfeld G. J Appl Physiol 95: 73-80, 2003; Belay T, Aviles H, Vance M, Fountain K, and Sonnenfeld G. J Allergy Clin Immunol 110: 262-268, 2002). This is a model of some of the aspects of spaceflight conditions, including lack of load bearing on hindlimbs and a fluid shift to the head. Active hexose correlated compound (AHCC), extracted from Basidiomycete mushrooms, has been shown to induce enhancement of immune responses, including enhanced natural killer activity. In the present study, AHCC was orally administered to mice to determine whether the treatment could decrease immunosuppression and mortality of mice maintained in the hindlimb-unloaded model and infected with Klebsiella pneumoniae. The results of the present study showed that administration of AHCC by gavage for 1 wk (1 g/kg body wt) before suspension and throughout the 10-day suspension period yielded significant beneficial effects for the hindlimb-unloaded group, including 1). decreased mortality, 2). increased time to death, and 3). increased ability to clear bacteria. The results suggest that AHCC can decrease the deleterious effects of the hindlimb-unloading model on immunity and resistance to infection.     

Polyribosome formation and protein synthesis in imbibed but dormant lettuce seeds

Dormancy is maintained in Grand Rapids lettuce (Lactuca sativa) seeds imbibed on water in darkness at 25 C. Polyribosome formation and protein synthesis occur early in the imbibition phase and considerable polysomal material is also present after 24 and 48 hours, even though the seeds have failed to germinate. Incorporation of labeled leucine into protein following a 24-hour preincubation period shows that these polysomes are active in protein synthesis.     

Distance,environmental and substrate factors impacting recovery of bryophyte communities after harvesting

Aims

Bryophyte re‐colonization after disturbance is largely governed by environmental conditions within disturbed forests. In particular, distance to a forest edge is an important predictor of bryophyte community re‐colonization, through either direct constraints, such as dispersal limitation, or indirectly by altering environmental conditions. This study examines a range of factors – environmental, distance to an edge, substrate specific environment or local‐level environment – to determine which are important in the re‐colonization of bryophyte communities after forest harvesting. As bryophyte communities vary with the particular substrate inhabited, responses were examined across four substrates (rock, exposed roots, ground and CWD).

Location

Tasmanian southern forests, Australia.

Methods

Bryophyte composition was examined on four substrates (ground, coarse wood debris, exposed roots, rocks) within three ages (~7, ~27 and ~45 years post‐disturbance) of harvested wet eucalypt forest. Re‐colonization success of bryophyte communities was determined by comparing communities in regeneration forest to mature forest communities using axis scores from one‐dimensional constrained ordination. The importance of various environmental conditions for re‐colonization success was then modelled. Finally, path analysis was used to determine whether the impact of distance to a forest edge was meditated through its effects on key environmental variables.

Results

Multiple environmental factors impacted re‐colonization of mature bryophyte communities. Local‐level conditions such as microclimate (temperature, humidity and VPD) and LAI were the most important in determining re‐colonization across substrates. Path analysis showed that distance to a forest edge had a significant impact on re‐colonization success, but only a relatively small part of this was mediated through its impact on environmental factors.

Conclusions

Bryophyte re‐colonization is driven by a combination of microclimate conditions and factors related to distance from a forest edge (most likely dispersal distance). While some substrate‐specific factors impact bryophyte re‐colonization success, the consistent impact of local environmental factors across substrates suggests that harvesting management strategies that develop more ‘mature’ microclimate conditions and increase proximity to nearby mature forest patches will be beneficial for all bryophytes communities. As bryophyte re‐colonization was correlated with temporally dynamic environmental conditions, we suggest that forest age needs to be considered in future work.     

Pulmonary aspiration in preschool children with cystic fibrosis

Pulmonary aspiration of gastric refluxate (PAGR) has been demonstrated in association with pulmonary inflammation in school aged children with Cystic Fibrosis (CF). We sought to determine if similar findings were present in preschool children. Pepsin was measured in Broncho-alveolar lavage (BAL) fluid collected from clinically stable preschool children with CF and controls. Elevated pepsin levels were found in a subgroup of children with CF, but this was not found to be associated with pulmonary infection, pulmonary inflammation or respiratory or gastrointestinal symptoms.     

P2Y<Subscript>2</Subscript> and P2Y<Subscript>6</Subscript> receptor activation elicits intracellular calcium responses in human adipose-derived mesenchymal stromal cells

Adipose tissue contains self-renewing multipotent cells termed mesenchymal stromal cells. In situ, these cells serve to expand adipose tissue by adipogenesis, but their multipotency has gained interest for use in tissue regeneration. Little is known regarding the repertoire of receptors expressed by adipose-derived mesenchymal stromal cells (AD-MSCs). The purpose of this study was to undertake a comprehensive analysis of purinergic receptor expression. Mesenchymal stromal cells were isolated from human subcutaneous adipose tissue and confirmed by flow cytometry. The expression profile of purinergic receptors was determined by quantitative real-time PCR and immunocytochemistry. The molecular basis for adenine and uracil nucleotide-evoked intracellular calcium responses was determined using Fura-2 measurements. All the known subtypes of P2X and P2Y receptors, excluding P2X2, P2X3 and P2Y₁₂ receptors, were detected at the mRNA and protein level. ATP, ADP and UTP elicited concentration-dependent calcium responses in mesenchymal cells (N?=?7–9 donors), with a potency ranking ADP (EC₅₀ 1.3 ± 1.0 μM)?>?ATP (EC₅₀ 2.2 ± 1.1 μM)?=?UTP (3.2 ± 2.8 μM). Cells were unresponsive to UDP (<?30 μM) and UDP-glucose (<?30 μM). ATP responses were attenuated by selective P2Y₂ receptor antagonism (AR-C118925XX; IC₅₀ 1.1 ± 0.8 μM, 73.0?±?8.5% max inhibition; N?=?7 donors), and UTP responses were abolished. ADP responses were attenuated by the selective P2Y₆ receptor antagonist, MRS2587 (IC₅₀ 437 ± 133nM, 81.0?±?8.4% max inhibition; N?=?6 donors). These data demonstrate that adenine and uracil nucleotides elicit intracellular calcium responses in human AD-MSCs with a predominant role for P2Y₂ and P2Y₆ receptor activation. This study furthers understanding about how human adipose-derived mesenchymal stromal cells can respond to external signalling cues.