The human amniotic fluid stem cell secretome triggers intracellular Ca(2+) oscillations, NF‐κB nuclear translocation and tube formation in human endothelial colony‐forming cells

Second trimester foetal human amniotic fluid‐derived stem cells (hAFS) have been shown to possess remarkable cardioprotective paracrine potential in different preclinical models of myocardial injury and drug‐induced cardiotoxicity. The hAFS secretome, namely the total soluble factors released by cells in their conditioned medium (hAFS‐CM), can also strongly sustain in vivo angiogenesis in a murine model of acute myocardial infarction (MI) and stimulates human endothelial colony‐forming cells (ECFCs), the only truly recognized endothelial progenitor, to form capillary‐like structures in vitro. Preliminary work demonstrated that the hypoxic hAFS secretome (hAFS‐CM(Hypo)) triggers intracellular Ca(2+) oscillations in human ECFCs, but the underlying mechanisms and the downstream Ca(2+)‐dependent effectors remain elusive. Herein, we found that the secretome obtained by hAFS undergoing hypoxic preconditioning induced intracellular Ca(2+) oscillations by promoting extracellular Ca(2+) entry through Transient Receptor Potential Vanilloid 4 (TRPV4). TRPV4‐mediated Ca(2+) entry, in turn, promoted the concerted interplay between inositol‐1,4,5‐trisphosphate‐ and nicotinic acid adenine dinucleotide phosphate‐induced endogenous Ca(2+) release and store‐operated Ca(2+) entry (SOCE). hAFS‐CM(Hypo)‐induced intracellular Ca(2+) oscillations resulted in the nuclear translocation of the Ca(2+)‐sensitive transcription factor p65 NF‐κB. Finally, inhibition of either intracellular Ca(2+) oscillations or NF‐κB activity prevented hAFS‐CM(Hypo)‐induced ECFC tube formation. These data shed novel light on the molecular mechanisms whereby hAFS‐CM(Hypo) induces angiogenesis, thus providing useful insights for future therapeutic strategies against ischaemic‐related myocardial injury.