Key result
In an intact perfused mouse lung model, Na/K-ATPase activity was completely inhibited by 1 mM ouabain, whereas oleic acid caused approximately half the inhibition of ouabain (p<0.002).
A radioactivity-free assay using ICP-OES successfully measured Na/K-ATPase activity in intact perfused mice lungs, demonstrating complete inhibition by ouabain and partial inhibition by oleic acid.
Validates ICP-OES assay for lung Na/K-ATPase in mice; leaves open translation to human models or clinical use.
BACKGROUND: Among the characteristics of acute respiratory distress syndrome (ARDS) is edema formation and its resolution depends on pneumocyte Na/K-ATPase activity. Increased concentration of oleic acid (OA) in plasma induces lung injury by targeting Na/K-ATPase and, thus, interfering in sodium transport. FINDINGS: Presently, we adapted a radioactivity-free assay to detect Na/K-ATPase activity in perfused lung mice, comparing the inhibitory effect of ouabain and OA. We managed to perfuse only the lung, avoiding the systemic loss of rubidium. Rb+ incorporation into lung was measured by inductively coupled plasma optical emission spectrometry (ICP OES) technique, after lung tissue digestion. Na/K-ATPase activity was the difference between Rb+ incorporation with or without ouabain. Lung Na/K-ATPase was completely inhibited by perfusion with ouabain. However, OA caused a partial inhibition. CONCLUSIONS: In the present work the amount of incorporated Rb+ was greater than seen in our previous report, showing that the present technique is trustworthy. This new proposed assay may allow researchers to study the importance of Na/K-ATPase activity in lung pathophysiology.
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Gonçalves-de-Albuquerque et al. (2014) studied Acute respiratory distress syndrome (ARDS) model. Ouabain and Oleic acid vs. Control (KCl free-Hank's solution) was evaluated on Rb+ incorporation in lung tissue (Na/K-ATPase activity) (p=<0.002). In an intact perfused mouse lung model, Na/K-ATPase activity was completely inhibited by 1 mM ouabain, whereas oleic acid caused approximately half the inhibition of ouabain (p<0.002).
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