An investigation of pressure ulcer risk, comfort and pain in medical imaging

In this study, we investigated the interface pressure of healthy volunteers on medical imaging (MI) table surfaces to determine the risks of developing pressure ulcers (PU). We also investigated volunteers’ perception of pain and comfort while lying on the MI table surfaces. Evidence from this study will enhance the understanding of factors contributing to PU formation and help improve service delivery to patients undergoing MI procedures

Effect of Fructooligosaccharide Metabolism on Chicken Colonization by an Extra-Intestinal Pathogenic Escherichia coli Strain

Extra-intestinal pathogenic Escherichia coli (ExPEC) strains cause many diseases in humans and animals. While remaining asymptomatic, they can colonize the intestine for subsequent extra-intestinal infection and dissemination in the environment. We have previously identified the fos locus, a gene cluster within a pathogenicity island of the avian ExPEC strain BEN2908, involved in the metabolism of short-chain fructooligosaccharides (scFOS). It is assumed that these sugars are metabolized by the probiotic bacteria of the microbiota present in the intestine, leading to a decrease in the pathogenic bacterial population. However, we have previously shown that scFOS metabolism helps BEN2908 to colonize the intestine, its reservoir. As the fos locus is located on a pathogenicity island, one aim of this study was to investigate a possible role of this locus in the virulence of the strain for chicken. We thus analysed fos gene expression in extracts of target organs of avian colibacillosis and performed a virulence assay in chickens. Moreover, in order to understand the involvement of the fos locus in intestinal colonization, we monitored the expression of fos genes and their implication in the growth ability of the strain in intestinal extracts of chicken. We also performed intestinal colonization assays in axenic and Specific Pathogen-Free (SPF) chickens. We demonstrated that the fos locus is not involved in the virulence of BEN2908 for chickens and is strongly involved in axenic chicken cecal colonization both in vitro and in vivo. However, even if the presence of a microbiota does not inhibit the growth advantage of BEN2908 in ceca in vitro, overall, growth of the strain is not favoured in the ceca of SPF chickens. These findings indicate that scFOS metabolism by an ExPEC strain can contribute to its fitness in ceca but this benefit is fully dependent on the bacteria present in the microbiota

The impact of immediate breast reconstruction on the time to delivery of adjuvant therapy: the iBRA-2 study

Background: Immediate breast reconstruction (IBR) is routinely offered to improve quality-of-life for women requiring mastectomy, but there are concerns that more complex surgery may delay adjuvant oncological treatments and compromise long-term outcomes. High-quality evidence is lacking. The iBRA-2 study aimed to investigate the impact of IBR on time to adjuvant therapy. Methods: Consecutive women undergoing mastectomy ± IBR for breast cancer July–December, 2016 were included. Patient demographics, operative, oncological and complication data were collected. Time from last definitive cancer surgery to first adjuvant treatment for patients undergoing mastectomy ± IBR were compared and risk factors associated with delays explored. Results: A total of 2540 patients were recruited from 76 centres; 1008 (39.7%) underwent IBR (implant-only [n = 675, 26.6%]; pedicled flaps [n = 105,4.1%] and free-flaps [n = 228, 8.9%]). Complications requiring re-admission or re-operation were significantly more common in patients undergoing IBR than those receiving mastectomy. Adjuvant chemotherapy or radiotherapy was required by 1235 (48.6%) patients. No clinically significant differences were seen in time to adjuvant therapy between patient groups but major complications irrespective of surgery received were significantly associated with treatment delays. Conclusions: IBR does not result in clinically significant delays to adjuvant therapy, but post-operative complications are associated with treatment delays. Strategies to minimise complications, including careful patient selection, are required to improve outcomes for patients

A new dimension to Ras function: a novel role for nucleotide-free Ras in Class II phosphatidylinositol 3-kinase beta (PI3KC2β) regulation.

The intersectin 1 (ITSN1) scaffold stimulates Ras activation on endocytic vesicles without activating classic Ras effectors. The identification of Class II phosphatidylinositol 3-kinase beta, PI3KC2β, as an ITSN1 target on vesicles and the presence of a Ras binding domain (RBD) in PI3KC2β suggests a role for Ras in PI3KC2β activation. Here, we demonstrate that nucleotide-free Ras negatively regulates PI3KC2β activity. PI3KC2β preferentially interacts in vivo with dominant-negative (DN) Ras, which possesses a low affinity for nucleotides. PI3KC2β interaction with DN Ras is disrupted by switch 1 domain mutations in Ras as well as RBD mutations in PI3KC2β. Using purified proteins, we demonstrate that the PI3KC2β-RBD directly binds nucleotide-free Ras in vitro and that this interaction is not disrupted by nucleotide addition. Finally, nucleotide-free Ras but not GTP-loaded Ras inhibits PI3KC2β lipid kinase activity in vitro. Our findings indicate that PI3KC2β interacts with and is regulated by nucleotide-free Ras. These data suggest a novel role for nucleotide-free Ras in cell signaling in which PI3KC2β stabilizes nucleotide-free Ras and that interaction of Ras and PI3KC2β mutually inhibit one another

A New Dimension to Ras Function: A Novel Role for Nucleotide-Free Ras in Class II Phosphatidylinositol 3-Kinase Beta (PI3KC2 beta) Regulation

The intersectin 1 (ITSN1) scaffold stimulates Ras activation on endocytic vesicles without activating classic Ras effectors. The identification of Class II phosphatidylinositol 3-kinase beta, PI3KC2 beta, as an ITSN1 target on vesicles and the presence of a Ras binding domain (RBD) in PI3KC2 beta suggests a role for Ras in PI3KC2 beta activation. Here, we demonstrate that nucleotide-free Ras negatively regulates PI3KC2 beta activity. PI3KC2 beta preferentially interacts in vivo with dominant-negative (DN) Ras, which possesses a low affinity for nucleotides. PI3KC2 beta interaction with DN Ras is disrupted by switch 1 domain mutations in Ras as well as RBD mutations in PI3KC2 beta. Using purified proteins, we demonstrate that the PI3KC2 beta-RBD directly binds nucleotide-free Ras in vitro and that this interaction is not disrupted by nucleotide addition. Finally, nucleotide-free Ras but not GTP-loaded Ras inhibits PI3KC2 beta lipid kinase activity in vitro. Our findings indicate that PI3KC2 beta interacts with and is regulated by nucleotide-free Ras. These data suggest a novel role for nucleotide-free Ras in cell signaling in which PI3KC2 beta stabilizes nucleotide-free Ras and that interaction of Ras and PI3KC2 beta mutually inhibit one another

Mutations in the effector region of Ras disrupt PI3KC2β binding.

<p>(A) Point mutations in the effector region of Ras12V disrupt interactions with specific Ras targets. (B) VC-tagged PI3KC2β was co-transfected with either VN-tagged Ras17N, 17N/69N, or one the effector mutants in the background of Ras17N/69N. BiFC signal is pseudo-colored green. Effector mutations that disrupt Class I PI3K binding to Ras12V disrupt Class II PI3K binding to Ras17N/69N. CFP (red) was used as a transfection control. (C) Graph represents the average fluorescence intensity per cell ± S.E.M. from at least three independent experiments. (*p = 0.02). (D) Western blot analysis demonstrates equal expression of all constructs. (E) Mutation of Thr392 to Asp or Lys379 to Ala in full-length PI3KC2β disrupts interaction with Ras17N. The ΔRBD mutant was also included as a negative control. Graph represents the average of three independent experiments (*p<0.05).</p

Biological activity of PI3KC2β versus Raf RBDs.

<p>(A) Expression of the Raf-RBD but not the PI3KC2β-RBD or GST alone inhibited EGF-stimulation of a Gal-Elk reporter assay <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0045360#pone.0045360-Adams1" target="_blank">[23]</a>. GST-RBDs were expressed equally. GST alone migrated at a faster rate and was not visible in this image. Results represent the average relative activation ± S.E.M. from at least three independent experiments. (*p<0.05 compared to unstimulated GST, **p<0.01 compared to EGF stimulated GST). (B). PI3KC2β-RBD dose-dependently inhibits the effect of Ras17N on Src-mediated transformation. NIH/3T3 cells were transfected with 100 ng of SrcY527F expression construct in the presence or absence of Ras17N. Co-expression of the PI3KC2β-RBD reverses the inhibitory effect of Ras17N on Src transformation whereas the Raf-RBD does not. In contrast, expression of the Raf-RBD alone, but not the PI3KC2β-RBD, significantly inhibited Src-mediated transformation. The results represent the average relative focus forming activity ± S.E.M. from three independent experiments performed in triplicate. Asterisks denote samples that were significantly different from Src alone (*p<0.05).</p

Ras forms a complex with PI3KC2β.

<p>(A) PI3KC2β preferentially interacts with Ras17N. VC-tagged PI3KC2β was co-transfected with one of the following VN-tagged Ras constructs: WT, 61L, 17N, or 17N/69N. BiFC signal (green) demonstrates that PI3KC2β interacted with Ras17N >17N/69N >WT >61L. CFP (red) was used as a transfection control. The graph represents the average fluorescence intensity per cell ± S.E.M. from at least three independent (*p<0.05). Western blot analysis demonstrates equivalent expression of all constructs. (B) PI3KC2β does not interact with Ras12V. VC-tagged PI3KC2β was co-transfected with one of the following VN-tagged Ras constructs: WT,17N, or 12V. BiFC signal (green) demonstrates that PI3KC2β interacted with Ras17N >WT >12V. CFP (red) was used as a transfection control. The graph represents the average fluorescence intensity per cell ± S.E.M. from at least three independent experiments (*p<0.05). Western blot analysis demonstrates expression of all constructs (size bar  = 20 μm).</p