First determination of ATP in alveolar epithelium in situ, effect of mesenchymal stem cells

Abstract

The ATP probe, GlnKII‐GFP (Perceval®; Berg, 2009) enables quantification of ATP in single cells. Here, we used the probe for the first time in situ to determine whether mitochondrial transfer from bone marrow‐derived mesenchymal stem cells (MSCs) increases ATP in pulmonary alveoli. We transfected mouse lungs with the probe, then 24 hours later we gave mice LPS by airway instillation, followed a further 4 hours later with instillation of MSCs. After another 24 hours, we isolated and blood‐perfused the lungs and viewed alveoli by live confocal microscopy. In control lungs, Perceval fluorescence was well evident in the alveolar epithelium (AE), indicating successful transfection. The fluorescence decreased by 25% after LPS (P<0.05), but was restored to control levels by MSC instillation (P<0.05). Concomitantly, induced surfactant secretion by alveolar type 2 cells, an ATP‐dependent response, was blocked by LPS (P<0.05), but restored by subsequent MSC instillation (P<0.05). We conclude, the Perceval probe enables ATP assays in intact cells of organ systems amenable to live confocal microscopy. Our findings indicate that in the AE, LPS decreased ATP and blocked surfactant secretion. However, MSCs rescued ATP production and ATP‐dependent surfactant secretion in this pneumonitis model. Thus, by restoring alveolar mitochondrial function, MSCs may rescue overall lung function in sepsis (support HL64896).