Identification of the Amino Acids 300–600 of IRS-2 as 14-3-3 Binding Region with the Importance of IGF-1/Insulin-Regulated Phosphorylation of Ser-573

Phosphorylation of insulin receptor substrate (IRS)-2 on tyrosine residues is a key event in IGF-1/insulin signaling and leads to activation of the PI 3-kinase and the Ras/MAPK pathway. Furthermore, phosphorylated serine/threonine residues on IRS-2 can induce 14-3-3 binding. In this study we searched IRS-2 for novel phosphorylation sites and investigated the interaction between IRS-2 and 14-3-3. Mass spectrometry identified a total of 24 serine/threonine residues on IRS-2 with 12 sites unique for IRS-2 while the other residues are conserved in IRS-1 and IRS-2. IGF-1 stimulation led to increased binding of 14-3-3 to IRS-2 in transfected HEK293 cells and this binding was prevented by inhibition of the PI 3-kinase pathway and an Akt/PKB inhibitor. Insulin-stimulated interaction between endogenous IRS-2 and 14-3-3 was observed in rat hepatoma cells and in mice liver after an acute insulin stimulus and refeeding. Using different IRS-2 fragments enabled localization of the IGF-1-dependent 14-3-3 binding region spanning amino acids 300–600. The 24 identified residues on IRS-2 included several 14-3-3 binding candidates in the region 300–600. Single alanine mutants of these candidates led to the identification of serine 573 as 14-3-3 binding site. A phospho-site specific antibody was generated to further characterize serine 573. IGF-1-dependent phosphorylation of serine 573 was reduced by inhibition of PI 3-kinase and Akt/PKB. A negative role of this phosphorylation site was implicated by the alanine mutant of serine 573 which led to enhanced phosphorylation of Akt/PKB in an IGF-1 time course experiment. To conclude, our data suggest a physiologically relevant role for IGF-1/insulin-dependent 14-3-3 binding to IRS-2 involving serine 573

The treatment experiences of people living with ileostomies: an interpretative phenomenological analysis

AIMS: To explore treatment and healthcare experiences of people living with ileostomies, so nurses can build on best practice whilst caring for these patients. Background Ileostomies, in which the small intestine is re-directed out of a stoma in the abdominal wall so waste is collected using a bag, are used to treat conditions including Inflammatory Bowel Disease. Existing research suggests that although life with a stoma can be challenging, ileostomy patients are largely supported by multi-disciplinary teams. However, more research is needed to understand the nuances of these relationships and experiences of treatment, with a view to improving clinical care. DESIGN: Participants took part in semi-structured interviews consisting of open ended questions about their experiences of life with an ileostomy and their treatments and time in hospital. Points of interest were followed up. Data were collected between July and December of 2014. METHODS: The qualitative method Interpretative Phenomenological Analysis was used to analyse interviews with 21 people living with ileostomies. FINDINGS: We present findings from two clinical themes: Issues around treatment and Relationships with multi-disciplinary teams. Surgical complications were common, and several participants reported concerns about addiction to painkillers. Varying attitudes were found around reversal surgery. Many reported experiencing distressing, dehumanising care, whilst some felt supported by excellent healthcare providers. CONCLUSION: People living with ileostomies have a very mixed experience of multi-disciplinary teams and treatments, and much could be done to support them more fully. For example, more information about ileostomies could be given to patients and more training given to nurses around stomas

Chronic Helicobacter pylori infection in a population in southern Sweden analysed by histopathology, immunoblot and ELISA serology.

Background. Many individuals are infected with the bacterium Helicobacter pylori. Some develop ulcers or mucosal atrophy. Aims. To correlate the histological characteristics of the H. pylori-induced gastritis to the immunoblot pattern of the H. pylori infection and to compare the presence of H. pylori bacteria in tissue specimens with ELISA serology and immunoblot analysis. Methods. One hundred and sixty-six consecutive patients were referred to gastroscopy. Forty patients were excluded for various reasons and 126 were included in the study. Results. Twenty-three patients had ulcerations and 25 erosions. Ninety-two (73%) had a chronic gastritis and in 90 (71%) it involved both the antrum and corpus. Ninety-one (72%), of whom 96% had a chronic gastritis, had visible bacteria in the tissue specimens, used as the 'gold standard' for the detection of infection. In patients with chronic gastritis 65 (70%) had positive H. pylori ELISA serology, 27 (30%) had negative H. pylori ELISA, while 76 (83%) had a positive immunoblot pattern. The ELISA positive patients had more advanced chronic gastritis but a lower frequency of metaplasia and atrophy. Acute inflammatory activity in the chronic gastritis had a high immunoreactivity to 120 kDa (CagA) protein and was significantly correlated to antibody reactivity to proteins in the 53-65 kDa range (heat shock proteins) and to a 43 kDa subunit. Metaplasia and atrophy in antrum was associated with a 62 kDa protein band. Conclusion. Almost all H. pylori-infected patients had a pangastritis, visible in both antrum and corpus. Acute inflammatory activity in the chronic gastritis and the presence of metaplasia and atrophy in antrum were associated with a specific immunoblot pattern, indicating infection with more virulent strains. Immunoblot analysis had a better sensitivity than ELISA H. pylori serology

Surface-associated metal catalyst enhances the sorption of perfluorooctanoic acid to multi-walled carbon nanotubes

Perfluorooctanoic acid (PFOA) sorption behaviour of two commercial multi-walled carbon nanotubes (MWCNTs) (C 150 P from Bayer MaterialScience: BA and C-MWNTs from NanoTechLabs Inc.: CP) was investigated from aqueous solution. The BA nanotubes contain Co/Mn/Mg/Al catalysts both on their outer surface and in the inner bore while CP contains Fe-based catalyst typically within the tubes. The adsorption isotherms of 14C-radiolabeled PFOA were measured by batch experiments and fitted to the Freundlich model (r20.92).. The adsorption affinity and capacity on BA were significantly higher than on CP. Increasing the pH reduced the adsorption of PFOA due to the electrostatic interaction between the pH-sensitive surface and the adsorbate. Increasing NaCl concentration lead to the aggregation of the MWCNTs reducing the available surface and thus the adsorption capacity. Removal of the catalyst from the outer surface of BA changed the electrophoretic mobility from positive to negative value also decreasing the adsorbed amount of PFOA. The surface charge of the surface-associated metal catalyst favors the electrostatic sorption of PFOA. Such surface modification may be a promising way to improve the sorption capacity of MWCNTs for pollutants like PFOA and to broaden their potential application in water purification