Influenza neuraminidase is delivered directly to the apical surface of MDCK cell monolayers

AbstractThe aim of this study was to investigate whether influenza neuraminidase travels directly from the Golgi complex to the apical domain of the plasma membrane in virally infected epithelial (MDCK) cell monolayers, or whether it passes transiently through the basolateral domain. Using a new assay for the delivery of neuraminidase to the plasma membrane, we found that the time course of transport of this protein from the Golgi complex to the apical surface of MDCK cell monolayers was very similar to that for influenza haemagglutinin, which is known to be delivered directly to its destination. In addition, a similar time course of neuraminidase transport was found in BHK cells, which are not asymmetric and in which delivery must therefore be direct. Finally, basolateral exposure of MDCK cell monolayers grown on nitro-cellulose filters to an anti-neuraminidase antibody was shown to have no effect on the delivery of active neuraminidase to the apical surface. We conclude from these results that neuraminidase, like haemagglutinin, is delivered directly to the apical surface

Clinical characteristics and treatment outcomes of women with heroin dependence in Johannesburg, South Africa

Background. There has been a steady increase in the number of women with heroin dependence in South Africa (SA). Data from developed countries suggest that women with substance use disorder have unique treatment needs. There are limited SA data on women with heroin dependence and their response to treatment.Objectives. To describe the clinical and psychosocial characteristics of women entering inpatient rehabilitation for heroin dependence, determine the outcomes of treatment 3 and 9 months after rehabilitation, and compare these findings with male heroin users.Methods. We conducted a longitudinal study of 44 women with heroin dependence who were admitted to a rehabilitation facility in the West Rand Municipality of Gauteng Province, SA. The participants were assessed during admission and 3 and 9 months after leaving inpatient rehabilitation. Structured interviews measured changes in drug use, psychopathology, social functioning, injecting and sexual behaviour, criminality and general health. Statistical analysis of these outcomes and comparison between women and men at 3 months and 9 months was performed by a generalised estimating equation. Fixed and time-varying covariates were included in the models.Results. At baseline, 40% of female participants were HIV-positive, 50% engaged in sex work, 27% were injecting heroin users, and 75% were diagnosed with a comorbid mental illness. Thirty-seven (84%) and 30 (68%) were re-interviewed at the 3- and 9-month follow-up points, respectively. Of these, 6 were abstinent from all substances at 3 months and 2 at 9 months. Compared with males, females had a higher prevalence of HIV infection (p=0.006) and mental illness (p=0.0002) at enrolment. At 9 months, women had similar levels of drug use and criminality to men but scored significantly worse in terms of general health, social function and risky sexual behaviour.Conclusions. Women with heroin dependence in Johannesburg have high rates of HIV infection and comorbid mental illness and low rates of abstinence after inpatient detoxification and psychosocial therapy. Women fared worse than men in many domains of treatment outcome. This study builds evidence for the need for gender-sensitive substance rehabilitation facilities in SA.

Use of isothermal microcalorimetry to monitor microbial activities

Isothermal calorimetry measures the heat flow of biological processes, which is proportional to the rate at which a given chemical or physical process takes place. Modern isothermal microcalorimeters make measurements of less than a microwatt of heat flow possible. As a result, as few as 10 000-100 000 active bacterial cells in culture are sufficient to produce a real-time signal dynamically related to the number of cells present and their activity. Specimens containing bacteria need little preparation, and isothermal microcalorimetry (IMC) is a nondestructive method. After IMC measurements, the undisturbed samples can be evaluated by any other means desired. In this review, we present a basic description of microcalorimetry and examples of microbiological applications of IMC for medical and environmental microbiology. In both fields, IMC has been used to quantify microbial activity over periods of hours or even days. Finally, the recent development of highly parallel instruments (up to 48 channels) and the constantly decreasing costs of equipment have made IMC increasingly attractive for microbiology. Miniaturization of isothermal calorimeters provides an even wider range of possibilitie

Engineering human cell-based, functionally integrated osteochondral grafts by biological bonding of engineered cartilage tissues to bony scaffolds

In this study, we aimed at developing and validating a technique for the engineering of osteochondral grafts based on the biological bonding of a chondral layer with a bony scaffold by cell-laid extracellular matrix. Osteochondral composites were generated by combining collagen-based matrices (Chondro-Gide) containing human chondrocytes with devitalized spongiosa cylinders (Tutobone) using a fibrin gel (Tisseel). We demonstrate that separate pre-culture of the chondral layer for 3 days prior to the generation of the composite allows for (i) more efficient cartilaginous matrix accumulation than no pre-culture, as assessed histologically and biochemically, and (ii) superior biological bonding to the bony scaffold than 14 days of pre-culture, as assessed using a peel-off mechanical test, developed to measure integration of bilayered materials. The presence of the bony scaffold induced an upregulation in the infiltrated cells of the osteoblast-related gene bone sialoprotein, indicative of the establishment of a gradient of cell phenotypes, but did not affect per se the quality of the cartilaginous matrix in the chondral layer. The described strategy to generate osteochondral plugs is simple to be implemented and--since it is based on clinically compliant cells and materials--is amenable to be readily tested in the clinic

Spin-Correlation Coefficients and Phase-Shift Analysis for p+3^3He Elastic Scattering

Angular Distributions for the target spin-dependent observables A$_{0y}$, A$_{xx}$, and A$_{yy}$ have been measured using polarized proton beams at several energies between 2 and 6 MeV and a spin-exchange optical pumping polarized $^3$He target. These measurements have been included in a global phase-shift analysis following that of George and Knutson, who reported two best-fit phase-shift solutions to the previous global p+$^3$He elastic scattering database below 12 MeV. These new measurements, along with measurements of cross-section and beam-analyzing power made over a similar energy range by Fisher \textit{et al.}, allowed a single, unique solution to be obtained. The new measurements and phase-shifts are compared with theoretical calculations using realistic nucleon-nucleon potential models.Comment: Submitted to Phys. Rev.

Expansion algorithm for the density matrix

A purification algorithm for expanding the single-particle density matrix in terms of the Hamiltonian operator is proposed. The scheme works with a predefined occupation and requires less than half the number of matrix-matrix multiplications compared to existing methods at low (90%) occupancy. The expansion can be used with a fixed chemical potential in which case it is an asymmetric generalization of and a substantial improvement over grand canonical McWeeny purification. It is shown that the computational complexity, measured as number of matrix multiplications, essentially is independent of system size even for metallic materials with a vanishing band gap.Comment: 5 pages, 4 figures, to appear in Phys. Rev.

Isothermal microcalorimetry provides new insights into biofilm variability and dynamics

The purpose of this study was to investigate a three-species in vitro biofilm with peri-implantitis-related bacteria for its variability and metabolic activity. Streptococcus sanguinis, Fusobacterium nucleatum, and Porphyromonas gingivalis were suspended in simulated body fluid containing 0.2% glucose to form biofilms on polished, protein-coated implant-grade titanium disks over 72 h using a flow chamber system. Thereafter, biofilm-coated disks were characterized by scanning electron microscopy and fluorescence in situ hybridization/confocal laser scanning microscopy. To assess metabolic activity within the biofilms, their heat flow was recorded for 480 h at 37 °C by IMC. The microscopic methods revealed that the total number of bacteria in the biofilms varied slightly among specimens (2.59 × 104 ± 0.67 × 104 cells mm−2), whereas all three species were found constantly with unchanged proportions (S. sanguinis 41.3 ± 4.8%, F. nucleatum 17.7 ± 2.1%, and P. gingivalis 41.0 ± 4.9%). IMC revealed minor differences in time-to-peak heat flow (20.6 ± 4.5 h), a trend consistent with the small variation in bacterial species proportions as shown by microscopy. Peak heat flow (35.8 ± 42.6 µW), mean heat flow (13.1 ± 22.0 µW), and total heat over 480 h (23.5 ± 37.2 J) showed very high variation. These IMC results may be attributed to differences in the initial cell counts and relative proportions of the three species, their distribution and embedment in exopolysaccharide matrix on the test specimens. The present results provide new insights into variability and dynamics of biofilms on titanium disks, aspects that should be explored in future studies of dental surface

Use of an isothermal microcalorimetry assay to characterize microbial oxalotrophic activity

Isothermal microcalorimetry (IMC) has been used in the past to monitor metabolic activities in living systems. A few studies have used it on ecological research. In this study, IMC was used to monitor oxalotrophic activity, a widespread bacterial metabolism found in the environment, and particularly in soils. Six model strains were inoculated in solid angle media with K-oxalate as the sole carbon source.Cupriavidus oxalaticus,Cupriavidus necator, andStreptomyces violaceoruber presented the highest activity (91, 40, and 55 μW, respectively) and a maximum growth rate (μmax h−1) of 0.264, 0.185, and 0.199, respectively, among the strains tested. These three strains were selected to test the incidence of different oxalate sources (Ca, Cu, and Fe-oxalate salts) in the metabolic activity. The highest activity was obtained in Ca-oxalate forC. oxalaticus. Similar experiments were carried out with a model soil to test whether this approach can be used to measure oxalotrophic activity in field samples. Although measuring oxalotrophic activity in a soil was challenging, there was a clear effect of the amendment with oxalate on the metabolic activity measured in soil. The correlation between heat flow and growth suggests that IMC analysis is a powerful method to monitor bacterial oxalotrophic activit