Role of Glutathione Redox State in Oxygen Sensing by Carotid Body Chemoreceptor Cells

Producción CientíficaThis article first presents some basic structural traits of the carotid body (CB) arterial chemoreceptors to understand the relationship between the arterial blood PO2 and the activation of chemoreceptor cells, which are the O2 sensing structures of the CB. Some considerations in relation to the intensity of CB blood flow and O2 consumption of the organ would allow us to define the threshold for the detection of the hypoxic stimulus, which would lead us to the cardinal theme of the article, namely whether at the PO2 levels detected by the CB there alterations in the genesis of re-active oxygen species (ROS). An alteration in the rate of ROS productionwould impinge on the glutathione system [reduced glutathione (GSH) and oxidized glutathione (GSSG)], causing modifications in the GSH/GSSG ratio that are detected by direct measurement; the GSH/GSSG system rep-resents the quantitatively most important mechanism to dispose ROS and to maintain the overall redox status or redox environment in mammalian cells.1 The relationship between GSH/GSSG and oxygen chemoreception is approached from two different points of view. We will measure GSH/GSSG levels and calculate the redox environment of the cells and correl-ation with the activity of chemoreceptor cells in normoxia and in hypoxia. We will also present data on pharmacological manipulation of the redox environment of the cells, as assessed by GSH/GSSG quotients, and pos-sible correlations with the level of activity of chemoreceptor cells. The possible mechanisms of coupling between ROS and the GSH/GSSG system to the cellular effector machineries have been reviewed.2,

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Nanoparticle tracking analysis monitors microvesicle and exosome secretion from immune cells.

Nanoparticle tracking analysis permits the determination of both the size distribution and relative concentration of microvesicles, including exosomes, in the supernatants of cultured cells and biological fluids. We have studied the release of microvesicles from the human lymphoblastoid T-cell lines Jurkat and CEM. Unstimulated, both cell lines release microvesicles in the size range 70-90 nm, which can be depleted from the supernatant by ultracentrifugation at 100 000 g, and by anti-CD45 magnetic beads, and which by immunoblotting also contain the exosome-associated proteins Alix and Tsg101. Incubation with known potentiators of exosome release, the ionophores monensin and A23187, resulted in a significant increase in microvesicle release that was both time and concentration dependent. Mass spectrometric analysis of proteins isolated from ultracentrifuged supernatants of A23187-treated cells revealed the presence of exosome-associated proteins including heat-shock protein 90, tubulin, elongation factor α1, actin and glyceraldehyde 3-phosphate dehydrogenase. Additionally, treatment of peripheral blood monocyte-derived dendritic cells with bacterial lipopolysaccharide displayed an increase in secreted microvesicles. Consequently, nanoparticle tracking analysis can be effectively applied to monitor microvesicle release from cells of the immune system

The desirable qualities of future doctors: a study of medical student perceptions

Background: There is a lack of consensus regarding the qualities possessed by the ideal doctor, and very limited research regarding the views of medical students on these qualities. Aims: To investigate the views of commencing medical students regarding the desirable qualities of doctors Methods: A survey containing a set of proposed desirable qualities of doctors identified from the existing literature was completed by 158 first year medical students. Results: The survey had a 75% response rate. Students rated the individual qualities of empathy, motivation to be a doctor, good verbal communication, ethically sound, integrity and honesty as the most important. A factor analysis identified six categories of qualities: Methodical Processing, Cognitive Capacity, People Skills, Generic Work Ethic, Role Certainty and Warmth. Significant differences in factor scores were found across subgroups of students (international and domestic students, with and without prior tertiary studies) on the following factors: Methodical Processing, which was scored highest by domestic students with prior tertiary studies, Cognitive Capacity, which was scored highest by domestic students without prior tertiary studies and Generic Work Ethic, which was scored highest by international students. Conclusions: Medical students identified a range of desirable personal qualities of a doctor which varied according to student characteristics, including their prior educational experience. Future research aiming to define such desirable qualities should include a broader range of stakeholders, including students at different training levels and institutions

Proteasome subunits encoded by the major histocompatibility complex are not essential for antigen presentation

Major histocompatibility complex (MHC) class I molecules bind and deliver peptides derived from endogenously synthesized proteins to the cell surface for survey by cytotoxic T lymphocytes. It is believed that endogenous antigens are generally degraded in the cytosol, the resulting peptides being translocated into the endoplasmic reticulum where they bind to MHC class I molecules. Transporters containing an ATP-binding cassette encoded by the MHC class II region seem to be responsible for this transport. Genes coding for two subunits of the '20S' proteasome (a multicatalytic proteinase) have been found in the vicinity of the two transporter genes in the MHC class II region, indicating that the proteasome could be the unknown proteolytic entity in the cytosol involved in the generation of MHC class I-binding peptides. By introducing rat genes encoding the MHC-linked transporters into a human cell line lacking both transporter and proteasome subunit genes, we show here that the MHC-encoded proteasome subunit are not essential for stable MHC class I surface expression, or for processing and presentation of antigenic peptides from influenza virus and an intracellular protei