Cow milk-derived exosomes enhance the activity of the Na+/K+ ATPase in Caco-2 cells via EP3 and EP4 receptors

The increase in bovine dairy consumption has been correlated with the development of many inflammatory diseases. Emerging evidence points to milk exosomes as potential causative agents. The cargo of exosome consists of cytokines that are elevated in inflammatory bowel disease and modulate the activity of the colonic Na+/K+ ATPase. Whether a cause effect relationship exists between the exosomes and the Na+/K+ ATPase is a question that has not been addressed before and constitutes the aim of this work, using Caco-2 cells as a model. Exosomes were extracted from skim milk by a series of ultra-centrifugations and filtrations. The activity of the pump was assayed by measuring the amount of inorganic phosphate liberated in presence and absence of ouabain. The exosomes increased dose and time-dependently the activity of the ATPase with a maximal effect observed at 1 h. This increase was mediated via PGE2 and still appeared in presence of a blocker of EP1 and EP2, was enhanced further when EP4 was blocked, and changed to an inhibition in presence of an EP3 antagonist, suggesting an involvement of both EP3 and EP4. The activation of the Na+/K + ATPase was significantly enhanced or reduced when cells were treated respectively with an inhibitor or an activator of PKA. Since EP4 and EP3 act via Gs and Gi respectively, they exert opposite effects on PKA and consequently on the ATPase. The stimulatory effect of EP3 on the ATPase predominates over that of EP4 resulting in a net activation of the pump.