Acoustically detectable cellular-level lung injury induced by fluid mechanical stresses in microfluidic airway systems

Huh et al. 10.1073/pnas.0610868104.

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Fig. 5. Controlled manipulation of plug length and propagation speed. For a given flow rate of liquid, the length of a liquid plug can be varied by changing the time interval over which air flow is shut off. (A) When liquid is injected at 10 ml/hr, blockage of air for ~1 s results in the formation of a 1-mm-long liquid plug. At the same liquid flow rate, shorter plugs having lengths of 330 mm (B) and 70 mm (C) can be generated by obstructing air flow for ~400 ms and ~100 ms, respectively. (Scale bars: 500 mm.) When the liquid flow rate is fixed, the speed of plug propagation is determined mainly by the flow rate of air. (D) A liquid plug progresses at ~13.9 mm/s when air flow is driven at ~9.4 liters/hr. (E) A smaller air flow rate (~7.1 liters/hr) leads to slower propagation (~3.6 mm/s). (Scale bars: 1 mm.)





SI Soundtrack 1

Soundtrack 1. Actual sounds of crackles generated by rupture of a liquid plug inside the airway microchannel.





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Immunostaining of Tight Junction Protein and Analysis. Tight junction formation during air-liquid interface (ALI) culture was confirmed by performing immunofluorescent staining of a transmembrane protein localized to tight junctions known as occludin. For staining, a confluent monolayer of small airway epithelial cells (SAECs) cultured at an ALI for 4 days was fixed in 99% ethanol for 30 min at 4°C. This step was followed by incubation in acetone at -20°C for 3 min. Subsequently, the cell layer was blocked with 3% BSA in PBS for 30 min and incubated with FITC-conjugated mouse anti-human occludin antibody solution (10 mg/ml in PBS) at 4°C overnight. Images of stained cells were taken at five different observation areas along the length of a microchannel. As a control condition, epithelial cells were cultured with continuous perfusion of culture media without the formation of an ALI in the upper chamber. Fluorescence intensity of the images of the cells cultured at an ALI was evaluated to be 108.3 ±17.1 (mean ± SD in arbitrary unit), whereas the intensity of control cells was found to be 14.1 ±5.4 (mean ± SD). The higher average intensity demonstrates the formation of occludin induced by ALI culture conditions.

This Article

  1. Published online before print November 15, 2007, doi: 10.1073/pnas.0610868104 PNAS November 27, 2007 vol. 104 no. 48 18886-18891
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