Lymphocyte subsets in healthy Malawians: Implications for immunologic assessment of HIV infection in Africa

BackgroundCD4+T lymphocyte measurements are the most important indicator of mortality in HIV-infected individuals in resource-limited settings. There is currently a lack of comprehensive immunophenotyping data from African populations to guide the immunologic assessment of HIV infection.ObjectiveTo quantify variation in absolute and relative lymphocyte subsets with age in healthy Malawians.MethodsLymphocyte subsets in peripheral blood of 539 healthy HIV-uninfected Malawians stratified by age were enumerated by flow cytometry.ResultsB and T–lymphocyte and T-lymphocyte subset absolute concentrations peaked in early childhood then decreased to adult levels, whereas lymphocyte subset proportions demonstrated much less variation with age. Adult lymphocyte subsets were similar to those in developed countries. In contrast, high B-lymphocyte and CD8+T-lymphocyte levels among children under 2 years, relative to those in developed countries, resulted in low CD4+T-lymphocyte percentages that varied little between 0 and 5 years (35% to 39%). The CD4+T-lymphocyte percentages in 35% of healthy children under 1 year and 18% of children age 1 to 3 years were below the World Health Organization threshold defining immunodeficiency in HIV-infected children in resource-limited settings. Thirteen percent of healthy children under 18 months old had a CD4:CD8T-lymphocyte ratio <1.0, which is commonly associated with HIV infection. All immunologic parameters except absolute natural killer lymphocyte concentration varied significantly with age, and percentage and overall absolute CD4+T-lymphocyte counts were higher in females than males.ConclusionAlthough lymphocyte subsets in Malawian adults are similar to those from developed countries, CD4+T-lymphocyte percentages in young children are comparatively low. These findings need to be considered when assessing the severity of HIV-related immunodeficiency in African children under 3 years

Practising the Space Between: Embodying Belief as an Evangelical Anglican Student

This article explores the formation of British evangelical university students as believers. Drawing on ethnographic fieldwork conducted with a conservative evangelical Anglican congregation in London, I describe how students in this church come to embody a highly cognitive, word-based mode of belief through particular material practices. As they learn to identify themselves as believers, practices of reflexivity and accountability enable them to develop a sense of narrative coherence in their lives that allows them to negotiate tensions that arise from their participation in church and broader social structures. I demonstrate that propositional belief – in contexts where it becomes an identity marker – is bound up with relational practices of belief, such that distinctions between “belief in” and “belief that” are necessarily blurred in the lives of young evangelicals

Sequence-Specific β-Peptide Synthesis by a Rotaxane-Based Molecular Machine

We report on the synthesis and operation of a three-barrier, rotaxane-based, artificial molecular machine capable of sequence-specific β-homo (β3) peptide synthesis. The machine utilizes nonproteinogenic β3-amino acids, a class of amino acids not generally accepted by the ribosome, particularly consecutively. Successful operation of the machine via native chemical ligation (NCL) demonstrates that even challenging 15- and 19-membered ligation transition states are suitable for information translation using this artificial molecular machine. The peptide-bond-forming catalyst region can be removed from the transcribed peptide by peptidases, artificial and biomachines working in concert to generate a product that cannot be made by either machine alone

Is Streptococcus pyogenes resistant or susceptible to Trimethoprim-Sulfamethoxazole?

Streptococcus pyogenes is commonly believed to be resistant to trimethoprim-sulfamethoxazole (SXT), resulting in reservations about using SXT for skin and soft tissue infections (SSTI) where S. pyogenes is involved. S. pyogenes\u27 in vitro susceptibility to SXT depends on the medium\u27s thymidine content. Thymidine allows S. pyogenes to bypass the sulfur-mediated inhibition of folate metabolism and, historically, has resulted in apparently reduced susceptibility of S. pyogenes to sulfur antibacterials. The low thymidine concentration in Mueller-Hinton agar (MHA) is now regulated. We explored S. pyogenes susceptibility to SXT on various media. Using two sets of 100 clinical S. pyogenes isolates, we tested for susceptibility using SXT Etests on MHA containing defibrinated horse blood and 20 mg/liter &beta;-NAD (MHF), MHA with sheep blood (MHS), MHA alone, MHA with horse blood (MHBA), and MHA with lysed horse blood (MHLHBA). European Committee on Antibacterial Susceptibility Testing (EUCAST) breakpoints defined susceptibility (MIC, &le;1 mg/liter) and resistance (MIC, &gt;2 mg/liter). In study 1, 99% of S. pyogenes isolates were susceptible to SXT on MHA, MHBA, and MHLHBA, with geometric mean MICs of 0.04, 0.04, and 0.05 mg/liter, respectively. In study 2, all 100 S. pyogenes isolates were susceptible to SXT on MHF, MHS, MHA, and MHLHBA with geometric mean MICs of 0.07, 0.16, 0.07, and 0.09 mg/liter, respectively. This study confirms the in vitro susceptibility of S. pyogenes to SXT, providing support for the use of SXT for SSTIs. A clinical trial using SXT for impetigo is ongoing

The decision-making process for senior cancer patients: treatment allocation of older women with operable breast cancer in the UK.

Objective: Up to 40% of women over 70 years with primary operable breast cancer in the UK are treated with primary endocrine therapy (PET) as an alternative to surgery. A variety of factors are important in determining treatment for older breast cancer patients. This study aimed to identify the patient and tumor factors associated with treatment allocation in this population. Methods: Prospectively collected data on treatment received (surgery vs. PET) were analysed with multivariable logistic regression using the variables age, modified Charlson Comorbidity Index (CCI), activities of daily living (ADL) score, Mini-Mental State Examination (MMSE) score, HER2 status, tumour size, grade and nodal status. Results: Data were available for 1,122 cancers in 1,098 patients recruited between February 2013 and June 2015 from 51 UK hospitals. About 78% of the population were treated surgically, with the remainder being treated with PET. Increasing patient age at diagnosis, increasing CCI score, large tumor size (5 cm or more) and dependence in one or more ADL categories were all strongly associated with non-surgical treatment (P<0.05). Conclusion: Increasing comorbidity, large tumor size and reduced functional ability are associated with reduced likelihood of surgical treatment of breast cancer in older patients. However, age itself remains a significant factor for non-surgical treatment; reinforcing the need for evidence-based guidelines

Proteomic analysis of NMDA receptor–adhesion protein signaling complexes

N-methyl-D-aspartate receptors (NMDAR) mediate long-lasting changes in synapse strength via downstream signaling pathways. We report proteomic characterization with mass spectrometry and immunoblotting of NMDAR multiprotein complexes (NRC) isolated from mouse brain. The NRC comprised 77 proteins organized into receptor, adaptor, signaling, cytoskeletal and novel proteins, of which 30 are implicated from binding studies and another 19 participate in NMDAR signaling. NMDAR and metabotropic glutamate receptor subtypes were linked to cadherins and L1 cell-adhesion molecules in complexes lacking AMPA receptors. These neurotransmitter– adhesion receptor complexes were bound to kinases, phosphatases, GTPase-activating proteins and Ras with effectors including MAPK pathway components. Several proteins were encoded by activity-dependent genes. Genetic or pharmacological interference with 15 NRC proteins impairs learning and with 22 proteins alters synaptic plasticity in rodents. Mutations in three human genes (NF1, Rsk-2, L1) are associated with learning impairments, indicating the NRC also participates in human cognition

Conserved in VivoPhosphorylation of Calnexin at Casein Kinase II Sites as Well as a Protein Kinase C/Proline-directed Kinase Site

Calnexin is a lectin-like chaperone of the endoplasmic reticulum (ER) that couples temporally and spatially N-linked oligosaccharide modifications with the productive folding of newly synthesized glycoproteins. Calnexin was originally identified as a major type I integral membrane protein substrate of kinase(s) associated with the ER. Casein kinase II (CK2) was subsequently identified as an ER-associated kinase responsible for the in vitro phosphorylation of calnexin in microsomes (Ou, W-J., Thomas, D. Y., Bell, A. W., and Bergeron, J. J. M. (1992) J. Biol. Chem. 267, 23789-23796). We now report on the in vivo sites of calnexin phosphorylation. After 32PO4 labeling of HepG2 and Madin-Darby canine kidney cells, immunoprecipitated calnexin was phosphorylated exclusively on serine residues. Using nonradiolabeled cells, we subjected calnexin immunoprecipitates to in gel tryptic digestion followed by nanoelectrospray mass spectrometry employing selective scans specific for detection of phosphorylated fragments. Mass analyses identified three phosphorylated sites in calnexin from either HepG2 or Madin-Darby canine kidney cells. The three sites were localized to the more carboxyl-terminal half of the cytosolic domain: S534DAE (CK2 motif), S544QEE (CK2 motif), and S563PR. We conclude that CK2 is a kinase that phosphorylates calnexin in vivo as well as in microsomes in vitro. Another yet to be identified kinase (protein kinase C and/or proline-directed kinase) is directed toward the most COOH-terminal serine residue. Elucidation of the signaling cascade responsible for calnexin phosphorylation at these sites in vivo may define a novel regulatory function for calnexin in cargo folding and transport to the ER exit sites

CK2 is a key regulator of SLC4A2-mediated Cl-/HCO3 (-) exchange in human airway epithelia

Transepithelial bicarbonate secretion by human airway submucosal glands and surface epithelial cells is crucial to maintain the pH-sensitive innate defence mechanisms of the lung. cAMP agonists stimulate HCO3 (-) secretion via coordinated increases in basolateral HCO3 (-) influx and accumulation, as well as CFTR-dependent HCO3 (-) efflux at the luminal membrane of airway epithelial cells. Here, we investigated the regulation of a basolateral located, DIDS-sensitive, Cl-/HCO3 (-) exchanger, anion exchanger 2 (AE2; SLC4A2) which is postulated to act as an acid loader, and therefore potential regulator of HCO3 (-) secretion, in human airway epithelial cells. Using intracellular pH measurements performed on Calu-3 cells, we demonstrate that the activity of the basolateral Cl-/HCO3 (-) exchanger was significantly downregulated by cAMP agonists, via a PKA-independent mechanism and also required Ca2+ and calmodulin under resting conditions. AE2 contains potential phosphorylation sites by a calmodulin substrate, protein kinase CK2, and we demonstrated that AE2 activity was reduced in the presence of CK2 inhibition. Moreover, CK2 inhibition abolished the activity of AE2 in primary human nasal epithelia. Studies performed on mouse AE2 transfected into HEK-293T cells confirmed almost identical Ca2+/calmodulin and CK2 regulation to that observed in Calu-3 and primary human nasal cells. Furthermore, mouse AE2 activity was reduced by genetic knockout of CK2, an effect which was rescued by exogenous CK2 expression. Together, these findings are the first to demonstrate that CK2 is a key regulator of Cl--dependent HCO3 (-) export at the serosal membrane of human airway epithelial cells

Nautilus at Risk – Estimating Population Size and Demography of Nautilus pompilius

The low fecundity, late maturity, long gestation and long life span of Nautilus suggest that this species is vulnerable to over-exploitation. Demand from the ornamental shell trade has contributed to their rapid decline in localized populations. More data from wild populations are needed to design management plans which ensure Nautilus persistence. We used a variety of techniques including capture-mark-recapture, baited remote underwater video systems, ultrasonic telemetry and remotely operated vehicles to estimate population size, growth rates, distribution and demographic characteristics of an unexploited Nautilus pompilius population at Osprey Reef (Coral Sea, Australia). We estimated a small and dispersed population of between 844 and 4467 individuals (14.6–77.4 km−2) dominated by males (83∶17 male∶female) and comprised of few juveniles (<10%).These results provide the first Nautilid population and density estimates which are essential elements for long-term management of populations via sustainable catch models. Results from baited remote underwater video systems provide confidence for their more widespread use to assess efficiently the size and density of exploited and unexploited Nautilus populations worldwide

Tyrosine Phosphorylation of Tau by the Src Family Kinases Lck and Fyn

<p>Abstract</p> <p>Background</p> <p>Tau protein is the principal component of the neurofibrillary tangles found in Alzheimer's disease, where it is hyperphosphorylated on serine and threonine residues, and recently phosphotyrosine has been demonstrated. The Src-family kinase Fyn has been linked circumstantially to the pathology of Alzheimer's disease, and shown to phosphorylate Tyr18. Recently another Src-family kinase, Lck, has been identified as a genetic risk factor for this disease.</p> <p>Results</p> <p>In this study we show that Lck is a tau kinase. <it>In vitro</it>, comparison of Lck and Fyn showed that while both kinases phosphorylated Tyr18 preferentially, Lck phosphorylated other tyrosines somewhat better than Fyn. In co-transfected COS-7 cells, mutating any one of the five tyrosines in tau to phenylalanine reduced the apparent level of tau tyrosine phosphorylation to 25-40% of that given by wild-type tau. Consistent with this, tau mutants with only one remaining tyrosine gave poor phosphorylation; however, Tyr18 was phosphorylated better than the others.</p> <p>Conclusions</p> <p>Fyn and Lck have subtle differences in their properties as tau kinases, and the phosphorylation of tau is one mechanism by which the genetic risk associated with Lck might be expressed pathogenically.</p