Aromatic Amino Acid Activation of Signaling Pathways in Bone Marrow Mesenchymal Stem Cells Depends on Oxygen Tension

The physiologic oxygen pressures inside the bone marrow environment are much lower than what is present in the peripheral circulation, ranging from 1-7%, compared to values as high as 10-13% in the arteries, lungs and liver. Thus, experiments done with bone marrow mesenchymal stem cells (BMMSCs) using standard culture conditions may not accurately reflect the true hypoxic bone marrow microenvironment. However, since aging is associated with an increased generation of reactive oxygen species, experiments done under 21%O-2 conditions may actually more closely resemble that of the aging bone marrow environment. Aromatic amino acids are known to be natural anti-oxidants. We have previously reported that aromatic amino acids are potent agonists for stimulating increases in intracellular calcium and phospho-c-Raf and in promoting BMMSC differentiation down the osteogenic pathway. Our previous experiments were performed under normoxic conditions. Thus, we next decided to compare a normoxic (21% O-2) vs. a hypoxic environment (3% O-2) alone or after treatment with aromatic amino acids. Reverse-phase protein arrays showed that 3% O-2 itself up-regulated proliferative pathways. Aromatic amino acids had no additional effect on signaling pathways under these conditions. However, under 21%O-2 conditions, aromatic amino acids could now significantly increase these proliferative pathways over this "normoxic'' baseline. Pharmacologic studies are consistent with the aromatic amino acids activating the extracellular calcium-sensing receptor. The effects of aromatic amino acids on BMMSC function in the 21% O-2 environment is consistent with a potential role for these amino acids in an aging environment as functional anti oxidants.