Journal of visualized experiments : JoVE
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Human macrophages are primary host cells of intracellular Mycobacterium tuberculosis (Mtb) infection and thus have a central role in immune control of tuberculosis (TB). We have established an experimental protocol to follow immune polarization of myeloid-derived cells into M1 (classically activated) or M2 (alternatively activated) macrophage-like cells through assessment with a 10-color flow cytometry panel that allows visualization and deep-characterization of green-fluorescent-protein (GFP)-labeled Mtb in diverse macrophages subsets. Monocytes obtained from healthy blood donors were polarized into M1 or M2 cells using differentiation with granulocyte macrophage-colony-stimulating factor (GM-CSF) or macrophage-colony stimulating factor (M-CSF) followed by polarization with IFN-γ and lipopolysaccharide (LPS) or IL-4, respectively. ⋯ Several M1/M2 markers were downregulated after Mtb infection, which suggests that Mtb can modulate macrophage polarization. In addition, 24 different cell clusters of different sizes were found to be uniquely distributed among the M1 and M2 uninfected and Mtb-infected cells at 24-hours post-infection. This M1/M2 flow cytometry protocol could be used as a backbone in Mtb-macrophage research and be adopted for special needs in different areas of research.
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Manipulation of fluids and particles by acoustic actuation at small scale is aiding the rapid growth of lab-on-a-chip applications. Megahertz-order surface acoustic wave (SAW) devices generate enormous accelerations on their surface, up to 108 m/s2, in turn responsible for many of the observed effects that have come to define acoustofluidics: acoustic streaming and acoustic radiation forces. These effects have been used for particle, cell, and fluid handling at the microscale-and even at the nanoscale. ⋯ Representative results for the electrode pattern deposited on the substrate as well as the performance of SAW generated on the surface are displayed in detail. Fabrication tricks and troubleshooting are covered as well. This procedure offers a practical protocol for high frequency SAW device fabrication and integration for future microfluidics applications.
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The unexpected problematic airway represents a large proportion of anesthesia-related morbidity and mortality. The retromolar or paraglossal approach is an alternative to the majority of the rigid instruments used for tracheal intubation, which follow the midline to access the glottis. This single-center, prospective case-series study offers an option to conventional laryngoscopy in case of a poor glottic view, introducing an instrument (the rigid tube for laryngoscopy) that uses the retromolar approach to accomplish tracheal intubation. ⋯ This tool was tested on 30 patients with an unsatisfactory glottic view when using the Macintosh laryngoscope and obtained excellent results with respect to intubation time and complications. The reduced visual field is the main limitation of this method, which requires a training period for reasonable expertise. This simple, robust, and cheap instrument could be a rescue option in case of a difficult airway.
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Traumatic brain injury (TBI) is a major cause of death and disability. Diffuse axonal injury (DAI) is the predominant mechanism of injury in a large percentage of TBI patients requiring hospitalization. ⋯ Historically, experimental models of DAI without focal injury have been difficult to design. Here we validate a simple, reproducible and reliable rodent model of DAI that causes widespread white matter damage without skull fractures or contusions.
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Pneumothorax (PTX) represents accumulation of the air in the pleural space. A large or tension pneumothorax can collapse the lung and cause hemodynamic compromise, a life-threatening disorder. ⋯ The use of lung US in diagnosis of PTX together with US-guided thoracentesis results in earlier and more precise management. The recommendations presented in this publication are aimed at improving the application of lung US in guiding neonatal PTX diagnosis and management.