Regulation of Human Mesenchymal Stem Cell Functions by an Autocrine Loop Involving NAD+ Release and P2Y11-Mediated Signaling

In several cell types, a regulated efflux of NAD+ across Connexin 43 hemichannels (Cx43 HC) can occur, and extracellular NAD+ (NAD+e) affects cell-specific functions. We studied the capability of bone marrow-derived human mesenchymal stem cells (MSC) to release intracellular NAD+ through Cx43 HC. NAD+ efflux, quantified by a sensitive enzymatic cycling assay, was significantly upregulated by low extracellular Ca2+ (5–6-fold), by shear stress (13-fold), and by inflammatory conditions (3.1- and 2.5-fold in cells incubated with lipopolysaccharide (LPS) or at 39°C, respectively), as compared with untreated cells, whereas it was downregulated in Cx43-siRNA-transfected MSC (by 53%) and by cell-to-cell contact (by 45%). Further, we show that NAD+e activates the purinergic receptor P2Y11 and a cyclic adenosin monophosphate (cAMP)/cyclic ADP-ribose/[Ca2+]i signaling cascade, involving the opening, unique to MSC, of L-type Ca2+ channels. Extracellular NAD+ enhanced nuclear translocation of cAMP/Ca2+-dependent trans...