A novel role of L-serine (L-Ser) for the expression of nuclear factor of activated T cells (NFAT)2 in receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclastogenesis in vitro

Multinucleated cell formation is crucial for osteoclastogenesis, and the expression of nuclear factor of activated T cells (NFAT)2 (NFATc1) is essential for this process. We previously found, using mouse RAW264 cells, that culture at high cell density blocked progression to the multinucleated cell stage induced by stimulation with receptor activator of nuclear factor kappaB ligand (RANKL). Here, we have confirmed this finding in a bone marrow cell system and extended the analysis further. A high cell density appeared to cause a change in the composition of the culture medium accompanying downregulation of NFAT2 expression, and we identified L-serine (LSer) as essential for the expression of NFAT2 induced by RANKL. Namely, culture at high cell density caused a depletion of LSer in the medium. Consequently, L-Ser appeared to exert its effect at an early stage under the regular conditions used for inducing the expression of c-Fos, an upstream regulator of NFAT2. D-Ser, an enantiomer of L-Ser, showed no NFAT2-inducing activity. The expression of NFAT2, using a retrovirus vector, could compensate for the depletion of L-Ser and resume the progression to the multinucleated cell stage. These results demonstrate a novel role for L-Ser in RANKL-induced osteoclastogenesis in vitro.