Fig. 1: In situ AT2 mitochondrial responses to hyperinflation.
a Confocal images of a live alveolus (alv) show AT2 (arrows) in a septum of the alveolar wall, as identified by Lysotracker Red (LTR) staining. Images show time dependent Ca2+ responses in the cytosol (cCa2+) and mitochondria (mCa2+) of selected AT2 to a single 15-second hyperinflation induced by increasing airway pressure from 5 to 15 cmH2O. Ca2+ dyes were given by alveolar microinfusion. Ca2+ increases are denoted by increases of pseudocolored gray levels. Images were obtained at airway pressure of 5 cmH2O. Scale bar, 10 µm. Repeat images were taken at 3 different locations per lung in 4 lungs per group. b Tracings from an experiment and group data show AT2 Ca2+ responses to hyperinflation (arrow). MCUF/F, mice floxed for the MCU; rtTaCreinactv, mice expressing rtTA-Cre not exposed to doxycycline; rtTaMCU−/−, mice lacking the MCU in the alveolar epithelium. Bars: mean ± SEM. n = 4 mice per group. Groups were compared using one-way ANOVA with Bonferroni correction. c Group data show hyperinflation-induced surfactant secretion as quantified by loss of AT2 fluorescence of the lamellar body dye, lysotracker red (LTR). Negligible fluorescence loss indicates negligible surfactant secretion. Bars: mean ± SEM. n = 4 lungs for each bar. Groups were compared using one-way ANOVA with Bonferroni correction. n.s., not significant. d, e Group data are determinations of alveolar hyperinflation-induced mitochondrial Ca2+ (d) and surfactant secretion (e) responses in the indicated groups. We crossed the MCUF/F mice with mice bearing an inducible Cre recombinase under the control of an SPC promoter (SPC-Cre-ERT2). Post-partum Cre activation by tamoxifen (i.p.) caused AT2 MCU deletion in these mice. ERT2MCU−/−, tamoxifen treated; ERT2Creinactv, tamoxifen untreated. Bars: mean ± SEM. n = 4 lungs for each bar, p-values are for two-tailed Student’s t-test.
