Timing, rather than the concentration of cyclic AMP, correlates to osteogenic differentiation of human mesenchymal stem cells

Previously, we demonstrated that protein kinase A (PKA) activation using dibutyryl-cAMP in human mesenchymal stem cells (hMSCs) induces in vitro osteogenesis and bone formation in vivo. To further investigate the physiological role of PKA in hMSC osteogenesis, we tested a selection of G-protein-coupled receptor ligands which signal via intracellular cAMP production and PKA activation. Treatment of hMSCs with parathyroid hormone, parathyroid hormone-related peptide, melatonin, epinephrine, calcitonin or calcitonin gene-related peptide did not result in accumulation of cAMP or induction of alkaline phosphatase (ALP) expression. The only ligand that did induce cAMP, prostaglandin E2, even inhibited ALP expression and mineralization, suggesting that physiological levels of cAMP may inhibit osteogenesis. Furthermore, intermittent exposure of hMSCs to dibutyryl-cAMP inhibited ALP expression, whereas we did not observe an inhibitive effect at low dibutyryl-cAMP concentrations. Taken together, our results demonstrate that cAMP can either stimulate or inhibit osteogenesis in hMSCs, depending on the duration, rather than the strength, of the signal provided.