Integrated Mucus Transport Activities of Airway Epithelia
The Marsico Lung Institute has extensive experience measuring the integrated pulmonary defense activity of mucus transport in both culture and small animal models. With respect to culture, we have long had in-house our “classic” system whereby cell cultures grown in 1.2 mm Transwell ALI culture systems develop spontaneous rotational mucus transport that can be quantitated by time-lapse fluorescence videomicroscopy (Fig. 13). These techniques have been supplemented recently with novel techniques designed to utilize in-house built culture systems to measure mucus transport rates in linear systems. The in vitro systems are complemented by a number of techniques to measure mucus transport in mouse models. These techniques include direct deposition of radiotracers or fluorescence tracers, followed by measurement of linear clearance rates with time in the trachea (Fig. 14) or nasopharynx (Fig. 15). In addition, techniques have been developed to deposit aerosols on pulmonary surfaces to measure whole lung mucus clearance, using external technetium radioisotope counting techniques (Fig. 16).
Figure 13. Time lapse fluorescence microscopy of air-liquid interface culture surface to measure mucus transport rates.
Figure 14. Mucociliary clearance in the murine posterior nasopharynx (soft palate) following aspiration of 6um fluorescent beads.
Figure 15. Mucociliary clearance (MCC) in the murine trachea following aspiration of fluorescent beads (6 um) in perflurocarbon (0.25 ul/g). The trachea of the anesthetized mouse is exposed and covered with water equilibrated mineral oil for imaging. The first images of MCC are obtained while the mouse is alive (the breathing motions are obvious) and then the anesthetized mouse is euthanized by exsanguination and MCC is again recorded. Images are obtained by placing the mouse under a fluorescent dissection microscope (Leica) and recording with a DAGE MTI video camera.
Figure 16. Time lapse fluorescence microscopy of air-liquid interface culture surface to measure mucus transport rates.